Antibodies binding HBA1c

Abstract

The present invention relates to antibodies or antigen binding fragments thereof binding to Hemoglobin A1c (HbA1c) and nucleic acids coding for at least a part of antibodies or antigen binding fragments thereof binding to Hemoglobin A1c (HbA1c) of the present invention. Furthermore, the present invention relates to the use of antibodies or antigen binding fragments thereof binding to Hemoglobin A1c (HbA1c), compositions or diagnostic compositions of the present invention for determining the amount or concentration of HbA1c in a sample and to methods of for determining the amount or concentration of HbA1c in a sample.

Description

[0001]

[0002] ANTIBODIES BINDING HBA1C

[0003] Field of the invention

[0004] The present invention relates to antibodies or antigen binding fragments thereof binding to Hemoglobin Ale (HbAlc) and nucleic acids coding for at least a part of antibodies or antigen binding fragments thereof of the present invention. Furthermore, the present invention relates to the use of antibodies or antigen binding fragments thereof binding to Hemoglobin Ale (HbAlc), compositions or diagnostic compositions of the present invention for determining the amount or concentration of HbAl c in a sample and to methods of determining the amount or concentration of HbAlc in a sample.

[0005] Background of the invention

[0006] Hemoglobin (Hb) is an oxygen-carrying, heme-containing protein, which is abundant in red blood cells and formed by the developing erythrocytes in bone marrow. It is a conjugated protein containing four heme groups having the property of reversible oxygenation. Each Hb molecule is able to bind four oxygen molecules and the main function is to transport oxygen and carbon dioxide (Roberts, W., Mcmillin, G., Burtis, C. & Bruns, D. in Tietz textbook of clinical chemistry and molecular diagnostics, 5th ed (Elsevier Inc, 2012)). The half-life of Hb is defined by erythrocyte survival time, which is relatively constant at around 120 days (Goldstein, D. E. et al. Tests of glycemia in diabetes. Diabetes Care 27, 1761-1773 (2004); Thomas, L. Clinical laboratory diagnostics: Use and assessment of clinical laboratory results, e-book (TH-books Verlagsgesellschaft, 2016)). Hb consists of a heterogeneous group of Hb subfractions and derivatives, including glycated hemoglobins (Roberts, W., Mcmillin, G., Burtis, C. & Bruns, D. in Tietz textbook of clinical chemistry and molecular diagnostics, 5th ed (Elsevier Inc, 2012)). The formation of glycated Hb is a normal part of the physiologic function cycle. The degree of glycation, apart from the half-life of erythrocytes, depends essentially on the degree as well as the duration of the blood glucose elevation (Thomas, L. Clinical laboratory diagnostics: Use and assessment of clinical laboratory results, e-book (TH-books Verlagsgesellschaft, 2016). During glycated hemoglobin formation, initially the N-terminal end of the β-chain hemoglobin and blood glucose, which passes through the erythrocyte cell membrane, interact through a non-enzymatic reaction to form an aldimine in a reversible reaction. In the second, irreversible reaction step towards glycated Hb, the aldimine undergoes an Amadori rearrangement to produce a stable ketoamine, namely glycohemoglobin (Sherwani, S. I., Khan, H. A., Ekhzaimy, A., Masood, A. & Sakharkar, M. K. Significance of HbAlc Test in Diagnosis and Prognosis of Diabetic Patients. Biomark Insights 11, 95-104, doi:10.4137 / BMI. S38440 (2016)). Total glycohemoglobin is subdivided into subfractions depending on the glycation sites and the reaction partners. The native nonglycated Hb is A0 (HbA0). The subfractions (HbAlal, HbAla2, HbAlb, and HbAlc) are the result of the glycation of the N-terminal amino acid valine of the Hb β-chain with different carbohydrates. The sum of these subfractions are called HbAl. HbAlc is formed by the exposure of the N-terminal amino acid valine of the Hb β-chain to plasma glucose and is the major fraction of glycohemoglobin (Goldstein, D. E. et al. Tests of glycemia in diabetes. Diabetes Care 27, 1761-1773 (2004); Thomas, L. Clinical laboratory diagnostics: Use and assessment of clinical laboratory results, e-book (TH-books Verlagsgesellschaft, 2016)).

[0007] Normal adult Hb consists predominantly of HbA (97%), which is made up of four polypeptide chains, two α- and two β-chains (α2β2) (Sherwani, S. L, Khan, H. A., Ekhzaimy, A., Masood, A. & Sakharkar, M. K. Significance of HbAlc Test in Diagnosis and Prognosis of Diabetic Patients. Biomark Insights 11, 95-104, doi: 10.4137 / BMI. S38440 (2016)). The remainder is composed of HbA2 (α2δ2; 2.5%) and fetal HbF (α2γ2; 0.5%). About 6% of total HbA is HbAl, comprised of HbAlal, HbAla2, HbAlb, and HbAlc fractions (defined by their electrophoretic and chromatographic properties) (Sherwani, S. I., Khan, H. A., Ekhzaimy, A., Masood, A. & Sakharkar, M. K. Significance of HbAlc Test in Diagnosis and Prognosis of Diabetic Patients. Biomark Insights 11, 95-104, (2016)). HbAlc is the most abundant of these fractions constituting approximately 80 % of the total HbAl fraction (Roberts, W., Mcmillin, G., Burtis, C. & Bruns, D. in Tietz textbook of clinical chemistry and molecular diagnostics, 5th ed (Elsevier Inc, 2012)).

[0008] Diabetes is characterized by chronic hyperglycemia and causes long-term complications like retinopathy, neuropathy and nephropathy. It generally accelerates macro and micro vascular changes. Due to lifestyle changes (i.e. unbalanced diet rich in carbohydrates and lipids and less exercise), diabetes has become a global epidemic. Historically, glucose measured in the fasting state (fasting plasma glucose [FPG] tests) or glucose measured two hours after a carbohydrate challenge (oral glucose tolerance tests [OGTTs]) have been the standard measures used to diagnose diabetes and to identify people at risk for diabetes (Juarez, D. T., Demaris, K. M., Goo, R., Mnatzaganian, C. L. & Wong Smith, H. Significance of HbAlc and its measurement in the diagnosis of diabetes mellitus: US experience Diabetes Metab Syndr Obes 7, 487-494, doi: 10.2147 / dmso. S39092 (2014)).

[0009] Long-term prospective studies, notably the Diabetes Control and Complications Trial (DCCT), the UK Prospective Diabetes Study (UKPDS), and the Epidemiology of Diabetes Interventions and Complications (EDIC) study have provided definite evidence that diabetic complications are directly related to mean glycemic value, as measured by the HbAlc concentration. The formation of HbAlc depends on the half-life of the erythrocytes and on the exposure to glucose (Goldstein, D. E. et al. Tests of glycemia in diabetes. Diabetes Care 27, 1761-1773 (2004); Calisti, L. & Tognetti, S. Measure of glycosylated hemoglobin. Acta Biomed 76 Suppl 3, 59-62 (2005)). Thus, a build-up of HbAlc within the erythrocyte reflects an estimate of the average level of glucose to which the erythrocyte has been exposed to (Thomas, L. Clinical laboratory diagnostics: Use and assessment of clinical laboratory results, e-book (TH-books Verlagsgesellschaft, 2016)). As the lifespan of red blood cells is approximately 120 days, HbAlc levels reflects average glycemia over the past two to three months, independent of day -to day glucose fluctuations, recent exercise or food ingestion (Roberts, W., Mcmillin, G., Burtis, C. & Bruns, D. in Tietz textbook of clinical chemistry and molecular diagnostics, 5th ed (Elsevier Inc, 2012); Parrinello, C. M. & Selvin, E. Beyond HbAlc and glucose: the role of nontraditional glycemic markers in diabetes diagnosis, prognosis, and management. Curr Diab Rep 14, 548, (2014)). This, in combination with a successful standardization, has made HbAlc a valuable diagnostic tool of diabetes, for example for evaluation of long-term glycemic control in diabetic patients and predicting risks for development and / or progression of diabetic complications. The world health organization (WHO), the American Diabetes Association (ADA), the European Association for the Study of Diabetes, Diabetes Canada, and the International Diabetes Federation have defined HbAlc of > 6.5 % (48 mmol / mol) as criteria for the diagnosis of diabetes. Individuals close to the 6.5 % diagnostic threshold (i.e. HbAlc > 6.0 % but < 6.5 %) should receive effective intervention, such as weight control measures (Sacks, D. B. in Tietz textbook of clinical chemistry and molecular diagnostics, 5th ed (eds C Burtis, E Ashwood, & D Bruns) (Elsevier Inc, 2012); AmericanDiabetesAssociation. Standards of Medical Care in Diabetes. Diabetes Care 41, S13-S27 (2018); Committee, D. C. C. P. G. E. Diabetes Canada 2018 Clinical Practice Guidelines for the Prevention and Management of Diabetes in Canada. Can J Diabetes 42, S1=S325 (2018); Organization, W. H. Use of glycated haemoglobin (HbAlc) in the diagnosis of diabetes mellitus: abbreviated report of a WHO Consultation, 2011); Ryden, L. et al. ESC Guidelines on diabetes, pre-diabetes, and cardiovascular diseases developed in collaboration with the EASD: the Task Force on diabetes, pre-diabetes, and cardiovascular diseases of the European Society of Cardiology (ESC) and developed in collaboration with the European Association for the Study of Diabetes (EASD). Eur Heart J 34, 3035-3087, doi:10.1093 / eurheartj / ehtl08 (2013); Weykamp, C. HbAlc: A Review of Analytical and Clinical Aspects. Ann Lab Med. 2013 Nov; 33(6): 393–400).

[0010] As previously described, during the formation of HbAlc, glucose attaches to the N-terminal of hemoglobin β-chains to form an unstable aldimine intermediate to then undergo an Amadori rearrangement to form a stable ketoamine. Thus, according to the international federation of clinical chemistry (IFCC) the definition of HbAlc as marker for diabetes diagnosis and monitoring is the substance fraction of the β chains of hemoglobin that have a stable hexose adduct on the N-terminal amino acid valine or β N-1-deoxyfructosyl-hemoglobin. By definition, glucose adducts or intermediate / labile forms of the binding to Hb are excluded.

[0011] Various methods have been established to test for HbAlc. The principle of all methods is to either separate glycated and non-glycated forms of hemoglobin or separate HbAlc specifically from other hemoglobins. This is accomplished by either differences in charge (ion-exchange HPLC) or structure (immunoassays / boronate affinity chromatography / enzymatic).

[0012] The Tina-quant® Hemoglobin Ale from Roche (see also EP 0329994 Al, EP 0559164 Al and EP 0598329 Al) is a liquid ready to use two component turbidimetric inhibition immunoassay (TINA) for the quantitative (quant) in vitro determination of hemoglobin Ale (HbAlc) in whole blood (WB) or hemolysate (HEM). An HbAlc-specifc antibody (Ab) binds sample derived native HbAlc, so a soluble antigen-antibody complex is formed. Native HbAlc competes with a synthetic polymer with multiple copies of the immunoreactive part of HbAlc (agglutinator) for the antibody, by which insoluble antibody-agglutinator complexes are formed. During measurement, insoluble complexes are detected via principals of turbidimetry. The higher the native HbAlc concentration, the lower the turbidity. By the Twin Test reaction mode not only HbAlc, but also total hemoglobin (Hb) is bichromatically determined in a single cuvette, allowing the determination of HbAlc in relation to the total Hb concentration present in one sample. In detail, the anticoagulated whole blood sample is hemolyzed using a detergent (Tetradecyltrimethylammonium bromide (TTAB))-containing reagent. Due to the cationic nature of TTAB, the liberated hemoglobin is rearranged increasing the accessibility of the HbAlc epitope. With the help of N-Methylisothiazolon (MIT), hemoglobin (Hb) is converted to the basic hematin having a characteristic absorption spectrum, which is bichromatically measured during the pre-incubation phase of the immunological reaction. Glycohemoglobin (HbAlc) reacts with the anti-HbAlc Ab to form soluble antigen-antibody complexes. The Ab binds to the glycated N-terminus of the β-chain of HbAlc. Since the specific HbAlc epitope is present only twice on the HbAlc molecule, insoluble complexes cannot form. Next, the agglutinator Polyhapten comprising the antigen to which the mAb binds is added, which reacts with the excess mAb to form insoluble antibody-Polyhapten complexes. These insoluble complexes can be measured turbidimetrically. The Twin Test reaction mode facilitates sequential measurements of Hb and HbAlc in a single cuvette. Thus, only one sample-pipetting step is required. The final ratio of HbAlc to Hb is expressed as mmol / mol according to IFCC or % HbAlc according to National Glycohemoglobin Standardization Program (NGSP).

[0013] The antibody used so far in turbidimetric inhibition immunoassays is a polyclonal antibody. Polyclonal antibodies have several disadvantages: in order to obtain polyclonal antibodies, one has to immunize and bleed animals any time a new batch is to be produced. However, besides the fact that animals have to be used for obtaining the antibodies, this also leads to variability between batches, which leads to huge efforts mitigating this variability in order to obtain similar and comparable results for each new batch. It also leads to larger costs due to the increased efforts and required animal care. A set of polyclonal antibodies recognizes a multitude of epitopes, which may cause cross-reactivity. However, it is also the cause for one big advantage of polyclonal antibodies, namely their usually higher sensitivity against an antigen compared to monoclonal antibodies (mAb). For example, M-3.51.56 or M-3.230.140 (EP 0 329 994 Al) are monoclonal antibodies that show an inferior performance in comparison to polyclonal and show a worse coefficient of variation in assays determining HbAlc levels in a sample, as shown herein below.

[0014] There is thus still a need to replace polyclonal antibodies with monoclonal antibodies, while having a similar usability in quantifying HbAl c in immunoassays, in particular having a similar coefficient of variation in an immunoassay and / or having a similar precision and / or repeatability in an immunoassay.

[0015] Summary of the invention

[0016] In a first aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof having at least 85 % sequence identity to SEQ ID NO: 2, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof having at least 85 % sequence identity to SEQ ID NO: 3, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof having at least 85 % sequence identity to SEQ ID NO: 4, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof having at least 85 % sequence identity to SEQ ID NO: 6.

[0017] The inventors have surprisingly found that the antibodies of the present invention can be used in immunoassays to determine the amount or concentration of HbAlc in a sample. As can be seen in the appended Examples the antibodies or antigen binding fragments thereof of the present invention show a surprisingly low coefficient of variation in immunoassays in comparison to other antibodies, in particular other monoclonal antibodies. It was particularly surprising to obtain the antibodies or antigen binding fragments of the present invention that have a similar coefficient of variation in immunoassays compared to polyclonal antibodies. As can be seen in FIG. 7 out of the more than 3000 clones obtained only two showed that low coefficient of variation and may be used to replace polyclonal antibodies for an immunoassay for determining the levels of HbAlc. Other monoclonal antibodies binding to HbAlc known in the art do not show such low coefficients of variation.

[0018] In a further aspect, the present invention relates to a nucleic acid coding for at least a part of an antibody or antigen binding fragment thereof of the present invention.

[0019] In another aspect, the present invention relates to a cell comprising an antibody or antigen binding fragment thereof, a nucleic acid, an expression constructor a vector of the present invention.

[0020] In a further aspect, the present invention relates to a composition comprising an antibody or antigen binding fragment thereof of the present invention and an acceptable carrier. In a further aspect, the present invention relates to a kit comprising an antibody or antigen binding fragment thereof, a composition or a diagnostic composition of the present invention.

[0021] In a further aspect, the present invention relates to a use of an antibody or antigen binding fragment thereof, a composition or a diagnostic composition of the present invention for determining the amount or concentration of HbAlc in a sample.

[0022] In another aspect, the present invention relates to a method of determining the levels of HbAlc in a sample comprising a step of bringing the sample into contact with an antibody or antigen binding fragment thereof of the present invention.

[0023] List of figures

[0024] FIG. 1A shows kinetic profiles for newly obtained antibodies binding to SA-grafted HbAlc peptide. Kinetic profiles of preselected antibodies obtained from the kinetic screening binding HbAlc peptide, concentration 150 nM, with Streptavidin mass load.

[0025] FIG. IB shows kinetic profiles for prior art antibodies binding to SA-grafted HbAlc peptide.

[0026] FIG. 2 shows kinetic profiles for antibodies of the present invention (A) and state of the art antibodies (B) binding HbAlc peptide at 37°C. Shown are series of increasing concentrations HbAlc-peptide (black), c = 11.1 - 270 nM, duplicates for concentration 30 nM overlayed by a Langmuir 1:1 fitting model, Rmax global, RI = 0 (grey).

[0027] FIG. 3 shows kinetic profiles for the antibody Fab-Fragment 55C4 binding monovalent to surface-displayed nat. protein HbAlc at 37°C. Shown is a series of increasing concentrations of the Fab fragment (black), c = 3.3 - 270 nM, duplicates for concentration 30 nM overlayed by a Langmuir 1:1 fitting model, Rmax global, RI = 0 (grey).

[0028] FIG. 4 shows a 2D ligand diagram representation of the binding of the 55C4 antibody to the HbAlc ligand. The ligand is shown as sticks while spheres represent the residues of the antibody paratope. Image generated with the MOE software (Molecular Operating Environment (MOE), 2020.09 Chemical Computing Group ULC, 1010 Sherbooke St. West, Suite #910, Montreal, QC, Canada, H3A 2R7, 2022.) FIG. 5 shows a 2D ligand diagram representation of the binding of the 65H2 antibody to the HbAlc ligand. The ligand is shown as sticks while spheres represent the residues of the antibody paratope. Image generated with the MOE software (Molecular Operating Environment (MOE), 2020.09 Chemical Computing Group ULC, 1010 Sherbooke St. West, Suite #910, Montreal, QC, Canada, H3A 2R7, 2022.)

[0029] FIG. 6 shows ELISA of 55C4 binding to rare HbAlc variants mutations Kamakura (L3V), Jabalpur (L3P), Benin City (T4P)and Würzburg (T4N) in comparison to the wild type (wt) peptide HbAl c(l -7)[Glu(BiPEG)-7]amid.

[0030] FIG. 7 shows the process of selecting antibodies binding to HbAlc.

[0031] FIG. 8 shows a sequence alignment of the light and heavy chain variable domain of 65H2, 54E11 and 55C4.

[0032] Detailed Description of the invention

[0033] Before the present invention is described in detail below, it is to be understood that this invention is not limited to the particular embodiments and examples described herein as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention which will be limited only by the appended claims. Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art.

[0034] Several documents are cited throughout the text of this specification. Each of the documents cited herein (including all patents, patent applications, scientific publications, manufacturer's specifications, instructions etc.), whether supra or infra, is hereby incorporated by reference in its entirety. In the event of a conflict between the definitions or teachings of such incorporated references and definitions or teachings recited in the present specification, the text of the present specification takes precedence.

[0035] In the following, the elements of the present invention will be described. These elements are listed with specific embodiments, however, it should be understood that they may be combined in any manner and in any number to create additional embodiments. The various described examples and preferred embodiments should not be construed to limit the present invention to only the explicitly described embodiments. This description should be understood to support and encompass embodiments which combine the explicitly described embodiments with any number of the disclosed and / or preferred elements. Furthermore, any permutations and combinations of all described elements in this application should be considered disclosed by the description of the present application unless the context indicates otherwise.

[0036] The following definitions and embodiments apply to the present disclosure in its entirety, especially to all aspects and embodiments of the invention.

[0037] Definitions

[0038] As used in this specification and the appended claims, the singular forms "a", "an", and "the include plural referents, unless the content clearly dictates otherwise.

[0039] The word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps. In some embodiments, the word “comprising” may include “consisting”.

[0040] The use of the alternative (e.g. “or”) should be understood to mean either one, both or any combination thereof of the alternatives.

[0041] The term “and / or” should be understood to mean either one or both of the alternatives.

[0042] Percentages, concentrations, amounts and other numerical data may be expressed or presented herein in a “range” format. It is to be understood that such a range format is used merely for convenience and brevity and thus should be interpreted flexibly to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub-range is explicitly recited. As an illustration, a numerical range of "4 % to 20 %" should be interpreted to include not only the explicitly recited values of 4 % to 20 %, but to also include individual values and sub-ranges within the indicated range. Thus, included in this numerical range are individual values such as 4, 5, 6, 7, 8, 9, 10,... 18, 19, 20 % and sub-ranges such as from 4-10 %, 5-15 %, 10-20 %, etc. This same principle applies to ranges reciting minimal or maximal values. Furthermore, such an interpretation should apply regardless of the breadth of the range or the characteristics being described.

[0043] As used herein and unless stated otherwise, it is to be understood that the term “about” is used synonymously with the term “approximately”. Illustratively and unless stated otherwise, the use of the term “about” when used in conjunction with a stated numerical value or range denotes somewhat more or somewhat less than the stated value or range, to within a range of ±15% of that stated, ±10% of that stated, ±5% of that stated, or conveniently ± 2% of that stated. Such values are thus encompassed by the scope of the claims reciting the terms “about” or “approximately”.

[0044] As used in the present disclosure, "% w / v" refers to weight by volume percent, which is a unit of concentration measuring the amount of solute in grams (g) expressed as a percent of the total volume of solution in milliliters (ml).

[0045] As used in the present disclosure, "% v / v" refers to volume by volume percent, which is a unit of concentration measuring the amount of a specific solute in milliliters (ml) expressed as a percent of the total volume of solution in milliliters (ml).

[0046] As used herein, the terms "room temperature" and "ambient temperature" are used interchangeably herein and refer to temperatures from at least about 15 °C, e.g. from about 15 °C to about 35 °C, from about 15 °C to about 30 °C, from about 15 °C to about 25 °C or from about 17 °C to about 22 °C. Such temperatures will include 15 °C, 16 °C, 17 °C, 18 °C, 19 °C, 20 °C, 21 °C and 22 °C.

[0047] The term "recombinant" in the context of the present disclosure means "made through genetic engineering". In some embodiments, a "recombinant protein" in the context of the present disclosure is not occurring naturally.

[0048] The term "naturally occurring" as used herein refers to the fact that an object can be found in nature. For example, a peptide or nucleic acid that is present in an organism (including viruses) and can be isolated from a source in nature and which has not been intentionally modified by man in the laboratory is naturally occurring. The term "found in nature" means "present in nature" and includes known objects as well as objects that have not yet been discovered and / or isolated from nature, but that may be discovered and / or isolated in the future from a natural source. The term “native” refers to a naturally occurring object.

[0049] According to the present invention, the term "peptide" refers to molecules which comprise about two or more, about 3 or more, about 4 or more, about 6 or more, about 8 or more, about 10 or more, about 13 or more, about 16 or more, about 20 or more, and up to about 50, about 100 or about 150, consecutive amino acids linked to one another via peptide bonds. The term "polypeptide" refers to large peptides, in particular peptides having at least about I51 amino acids. " Peptides" and "polypeptides" are both protein molecules. Thus, the terms "peptide", "protein" and "polypeptide" are used herein usually as synonyms.

[0050] For the purposes of the present invention, "variants" of an amino acid sequence (peptide or polypeptide) may comprise amino acid insertion variants, amino acid addition variants, amino acid deletion variants and / or amino acid substitution variants. The term "variant" includes all mutants, splice variants, post-translationally modified variants, conformations, isoforms, allelic variants, species variants, and species homologs, in particular those which are naturally occurring. The term "variant" includes, in particular, fragments of an amino acid sequence. Amino acid insertion variants comprise insertions of single or two or more amino acids in a particular amino acid sequence. In the case of amino acid sequence variants having an insertion, one or more amino acid residues are inserted into a particular site in an amino acid sequence, although random insertion with appropriate screening of the resulting product is also possible. Amino acid addition variants comprise amino- and / or carboxy-terminal fusions of one or more amino acids, such as 1, 2, 3, 5, 10, 20, 30, 50, or more amino acids. Amino acid deletion variants are characterized by the removal of one or more amino acids from the sequence, such as by removal of 1, 2, 3, 5, 10, 20, 30, 50, or more amino acids. The deletions may be in any position of the protein. Amino acid deletion variants that comprise the deletion at the N-terminal and / or C-terminal end of the protein are also called N-terminal and / or C- terminal truncation variants. Amino acid substitution variants are characterized by at least one residue in the sequence being removed and another residue being inserted in its place. Preference is given to the modifications being in positions in the amino acid sequence which are not conserved between homologous peptides or polypeptides and / or to replacing amino acids with other ones having similar properties. In some embodiments, amino acid changes in peptide and polypeptide variants are conservative amino acid changes, i.e. substitutions of similarly charged or uncharged amino acids. A conservative amino acid change involves substitution of one of a family of amino acids which are related in their side chains. Naturally occurring amino acids are generally divided into four families: acidic (aspartate, glutamate), basic (lysine, arginine, histidine), non-polar (alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine, tryptophan) and uncharged polar (glycine, asparagine, glutamine, cysteine, serine, threonine, tyrosine) amino acids. Phenylalanine, tryptophan, and tyrosine are sometimes classified jointly as aromatic amino acids. In some embodiments, conservative amino acid substitutions include substitutions within the following groups: glycine, alanine; valine, isoleucine, leucine; aspartic acid, glutamic acid; asparagine, glutamine; serine, threonine; lysine, arginine; and phenylalanine, tyrosine.

[0051] " Sequence identity" between two amino acid sequences indicates the percentage of amino acids that are identical between the sequences. " Sequence identity" between two nucleic acid sequences indicates the percentage of nucleotides that are identical between the sequences. The alignment for determining sequence similarity, such as sequence identity can be done with art known tools, such as using the best sequence alignment, for example, using Align, using standard settings, preferably EMBOSS::needle, Matrix: Blosum62, Gap Open 10.0, Gap Extend 0.5.

[0052] The terms "% identical" and "% identity" or similar terms are intended to refer, in particular, to the percentage of nucleotides or amino acids which are identical in an optimal alignment between the sequences to be compared. Said percentage is purely statistical, and the differences between the two sequences may be but are not necessarily randomly distributed over the entire length of the sequences to be compared. Comparisons of two sequences are usually carried out by comparing the sequences, after optimal alignment, with respect to a segment or "window of comparison", in order to identify local regions of corresponding sequences. The optimal alignment for a comparison may be carried out manually or with the aid of algorithms, e.g. the local homology algorithm by Smith and Waterman, 1981, Ads App. Math. 2, 482, the local homology algorithm by Needleman and Wunsch, 1970, J. Mol. Biol. 48, 443, the similarity search algorithm by Pearson and Lipman, 1988, Proc. Natl Acad. Sci. USA 88, 2444, or with the aid of computer programs using said algorithms (GAP, BESTFIT, FASTA, BLAST P, BLAST N and TFASTA in Wisconsin Genetics Software Package, Genetics Computer Group, 575 Science Drive, Madison, Wis.). In some embodiments, percent identity of two sequences is determined using the BLASTN or BLASTP algorithm, as available on the United States National Center for Biotechnology Information (NCBI) website. In some embodiments, the algorithm parameters used for BLASTN algorithm on the NCBI website include: (I) Expect Threshold set to 10; (II) Word Size set to 28; (III) Max matches in a query range set to 0; (IV) Match / Mismatch Scores set to 1, -2; (V) Gap Costs set to Linear; and (VI) the filter for low complexity regions being used. In some embodiments, the algorithm parameters used for BLASTP algorithm on the NCBI website include: (I) Expect Threshold set to 10; (II) Word Size set to 3; (III) Max matches in a query range set to 0; (IV) Matrix set to BLOSUM62; (V) Gap Costs set to Existence: 11 Extension: 1; and (VI) conditional compositional score matrix adjustment.

[0053] Percentage identity is obtained by determining the number of identical positions at which the sequences to be compared correspond, dividing this number by the number of positions compared (e.g. the number of positions in the reference sequence) and multiplying this result by 100.

[0054] The amino acid sequence variants described herein may readily be prepared by the skilled person, for example, by recombinant DNA manipulation. The manipulation of DNA sequences for preparing peptides or polypeptides having substitutions, additions, insertions or deletions, is described in detail in Molecular Cloning: A Laboratory Manual, 4th Edition, M. R. Green and J. Sambrook et al. (1989), eds., Cold Spring Harbor Laboratory Press, Cold Spring Harbor 2012, for example. Furthermore, the peptides, polypeptides and amino acid variants described herein may be readily prepared with the aid of known peptide synthesis techniques such as, for example, by solid phase synthesis and similar methods. In some embodiments, a fragment or variant of an amino acid sequence (peptide or polypeptide) is a "functional fragment" or "functional variant". The term "functional fragment" or "functional variant" of an amino acid sequence relates to any fragment or variant exhibiting one or more functional properties identical or similar to those of the amino acid sequence from which it is derived, i.e. it is functionally equivalent. With respect to sequences of binding agents such as antibodies, one particular function is one or more binding activities displayed by the amino acid sequence from which the fragment or variant is derived. The term "functional fragment" or "functional variant", as used herein, in particular refers to a variant molecule or sequence that comprises an amino acid sequence that is altered by one or more amino acids compared to the amino acid sequence of the parent molecule or sequence and that is still capable of fulfilling one or more of the functions of the parent molecule or sequence, e.g. binding to a target molecule. In some embodiments, the modifications in the amino acid sequence of the parent molecule or sequence do not significantly affect or alter the characteristics of the molecule or sequence. In different embodiments, the function of the functional fragment or functional variant may be reduced but still significantly present, e.g. function of the functional fragment or functional variant may be at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the parent molecule or sequence. However, in other embodiments, function of the functional fragment or functional variant may be enhanced compared to the parent molecule or sequence.

[0055] According to various embodiments of the present disclosure, a nucleic acid encoding a peptide or polypeptide is taken up by or introduced, i.e. transfected or transduced, into a cell which cell may be present in vitro or in a subject, resulting in expression of said peptide or polypeptide. The cell may, e.g. express the encoded peptide or polypeptide intracellularly (e.g. in the cytoplasm and / or in the nucleus), may secrete the encoded peptide or polypeptide, and / or may express it on the surface. According to the present disclosure, terms such as "nucleic acid expressing" and "nucleic acid encoding" or similar terms are used interchangeably herein and with respect to a particular peptide or polypeptide mean that the nucleic acid, if present in the appropriate environment, e.g. within a cell, can be expressed to produce said peptide or polypeptide.

[0056] The term "expression" as used herein includes the transcription and / or translation of a particular nucleotide sequence.

[0057] In the context of the present disclosure, the term "transcription" relates to a process, wherein the genetic code in a DNA sequence is transcribed into RNA (especially mRNA). Subsequently, the RNA may be translated into peptide or polypeptide.

[0058] With respect to RNA, the term "expression" or "translation" relates to the process in the ribosomes of a cell by which a strand of mRNA directs the assembly of a sequence of amino acids to make a peptide or polypeptide.

[0059] The term " C-terminus" refers to the carboxyl-terminus of a peptide chain. It is also called “carboxy-terminus” or “COOH-terminus”. A peptide chain comprises amino acids connected to one another via peptide bonds involving the amine group at the Caof the amino acid and the carboxyl-group. This forms a chain of amino acids connected via the amine group of X+1 amino acid and carboxyl-group of the X amino acid. As the amino acids at both ends are not further connected to another amino acid this leaves the amine group at one end and the carboxyl group at the other end free. The C-terminus is considered to be the end of the amino acid chain and the N-terminus the start of the chain. Amino acid sequences are therefore denoted starting from the amino acid with the free amine group at Cato the amino acid with the free carboxyl group at the Ca.

[0060] The term " N-terminus" refers to amine-terminus of a peptide chain. It is also called “aminoterminus”, “amine-terminus” or “NH2-terminus”. The N-terminus is considered to be the start of the amino acid chain.

[0061] The term “canonical amino acid” refers to any of the 20 amino acids used in general for polymerization with a ribosome, in particular the term refers to alanine, arginine, asparagine, aspartate, cysteine, glutamine, glutamate, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine.

[0062] The term "tag" refers to an amino acid sequence that is fused with a protein recombinantly. Tags can be used for various purposes such as purifying a protein or detected it in situ. Some tags can even serve two different purposes, such as maltose binding protein-tag, which can be used as a solubilisation enhancer and as an affinity tag for purification. Tags can be fused to a peptide or protein at the N-terminus or C-terminus or can be located within an amino acid sequence. A protein can have more than one tag, e.g. two affinity tags for two different purification steps or for different purposes such as detection and purification. The terms "tag", "protein tag" and "peptide tag" are used herein as synonyms. Examples of protein tags include His-tag, FLAG-tag, HA(hemagglutinin)-tag, Avi-tag, strep-tag, MBP(maltose binding protein)-tag, Fc-tag, GFP(green fluorescent protein)-tag or GST(glutatione S transferase)-tag.

[0063] The term "fusion protein" refers to a polypeptide or protein comprising two or more subunits. Preferably, the fusion protein is a translational fusion between the two or more subunits. The translational fusion may be generated by genetically engineering the coding nucleotide sequence for one subunit in a reading frame with the coding nucleotide sequence of a further subunit. Subunits may be interspersed by a linker.

[0064] The term "expression construct" refers to a nucleic acid comprising a coding sequence and further nucleotide sequences operatively linked to the coding sequence that are needed for the expression of the coding sequence. While a promoter and a terminator are required for efficiently starting and terminating translation of the coding sequence into mRNA, other sequences, such as 5' or 3' UTRs or enhancers are optional, but can aid expression of the coding sequence.

[0065] The term "isolated" means a protein or polypeptide, nucleic acid, cell, or other specified material or component that has been separated from at least one other material or component, including but not limited to, other proteins, nucleic acids, cells, etc. An isolated protein or polypeptide, nucleic acid, cell or other specified material or component may typically be in a form that does not occur in nature. For example, an isolated nucleic acid may be a nucleic acid that has been isolated from a cell, in which it originally was present.

[0066] The amount, concentration or level of a protein in a composition, such as a solution, may be measured by different methods known to the skilled person. Such methods may include mass spectrometry, UV absorbance, Bradford assay, Lowry assay or an immuno assay, such as ELISA. The percentage of a specific reconstituted protein from a lyophilisate may be determined by measuring the amount or concentration of the protein in the solution obtained by reconstitution and comparing this amount or concentration with the amount or concentration of the protein in the solution used to obtain the lyophilisate. When using concentration, the solution used to obtain the lyophilisate and the solution obtained by reconstitution have to have the same volume or the concentration values have to be adjusted to the same volume. For example, a lyophilisate X was produced using 500 µL of a solution comprising 2 mg / mL of protein Y. The lyophilisate was reconstituted to 1 mL and the concentration of protein Y in the resolubilized solution was 0.5 mg / mL. The adjusted concentration in the resolubilized solution is 1 mg / mL and the percentage of reconstituted protein is 50 % = (1 mg / mL ÷ 2 mg / mL) × 100.

[0067] The terms “antibody”, “antibodies”, and analogous terms as used herein relate to full immunoglobulin molecules and encompass naturally- occurring forms of antibodies (including but not limited to IgG, IgA, IgM, IgE) as well as recombinant antibody constructs including but not limited to single-chain antibodies, chimeric antibodies, humanized antibodies, antibodyfusion proteins and multi-specific antibodies, including but not limited to natural single domain antibodies (also referenced in the art as sdAbs and / or dAbs). Naturally occurring forms of antibodies generally comprise at least one pair of of VH and VL, each variable domain comprising three CDRs. The terms “antigen binding fragment” or “antigen binding fragments thereof’, which may be referenced herein as antibody antigen binding fragment, and / or, simply antigen binding fragment. These terms refer to one or more fragments of an antibody that retain the ability to specifically bind to the target antigen, i.e. HbAlc, as known in the art, including but not limited to antigen binding fragments comprising an Fv domain, i.e., paired heavy and light chain variable domains, such as Fab, Fab’, F(ab’)2, and Fv fragments as well as recombinant constructs such as singlechain Fv domains, known in the art as scFvs. The terms also includes antibody antigen binding fragments that comprise a single, unpaired heavy or light chain variable domain as known in the art that retains the ability to specifically and selectively bind antigen as defined herein, such as VHH domains based on the heavy chains of camelids (also known in the art as nanobodies).

[0068] In certain embodiments the antibody of the invention may be a full-immunoglobulin, Fab, Fab’, F(ab’)2, Fv, scFvor a protein comprising (VL-CL)nand (VH-CHl)n, wherein n is an integer of 2 to 12 and wherein each (VL-CL) or each (VH-CH1) is connected to the next (VL-CL) or next (VH-CH1) via a peptide linker. In a specific embodiment, the antibody of the invention may be a Fab fragment.

[0069] Antibodies may be polyclonal or monoclonal. The antibodies of the invention are monoclonal. The term “monoclonal” as used herein with reference to an antibody or antigen binding fragment thereof, refers to a population of antibody polypeptides or fragments thereof produced from a single B cell clone, which population contains only one species of an antigen binding site capable of immunoreacting with a particular epitope of an antigen. This is in contrast with “polyclonal” antibodies and compositions, which term(s) refer to a population of antibody polypeptides or antigen binding fragments that contain multiple species of antigen binding sites. Also included are modified forms of monoclonal antibodies of the invention such as humanized or chimeric versions thereof, as well as recombinant antibody constructs, such as antibody (or antigen binding fragment)-fusion proteins, wherein the antibody or antigen binding fragment comprises (an) additional domain(s), e.g. for the isolation and / or preparation of recombinantly produced antibody / fragment / constructs.

[0070] The term “variable region” or “variable domain” refers to the domain of an antibody heavy or light chain that is involved in binding the antibody to antigen. The variable domains of the heavy chain and light chain (VH and VL, respectively) of a native antibody generally have similar structures, with each domain comprising four conserved framework regions (FRs) and three complementary determining regions (CDRs). (See, e.g., Kindt et al. Kuby Immunology, 6th ed., W. H. Freeman and Co., page 91 (2007).) A single VH or VL domain may be sufficient to confer antigen-binding specificity. Furthermore, antibodies that bind a particular antigen may be isolated using a VH or VL domain from an antibody that binds the antigen to screen a library of complementary VL or VH domains, respectively. See, e.g., Portolano etal., J. Immunol. 150:880-887 (1993); Clarkson et al., Nature 352:624-628 (1991).

[0071] The term “paratope” as used herein in the context of antibodies relates to the amino acids of the VH and VL that directly interact with the antigen (e,g. HbAlc). Paratope amino acids may interact with the antigen via different interaction modes. Exemplary interaction modes are hydrophobic interaction, H-bond, H-bond with H2O coordination, and H2O coordination. The interaction of each paratope residue with the antigen may individually be mediated via the amino acid side chain or the amino acid backbone.

[0072] The term “hypervariable region” or “HVR” as used herein refers to each of the regions of an antibody variable domain which are hypervariable in sequence and which determine antigen binding specificity, for example “complementarity determining regions” (“CDRs”).

[0073] Generally, antibodies comprise six CDRs: three in the VH (CDR-H1, CDR-H2, CDR-H3), and three in the VL (CDR-L1, CDR-L2, CDR-L3). Exemplary CDRs herein include:

[0074] (a) hypervariable loops occurring at amino acid residues 26-32 (LI), 50-52 (L2), 91-96 (L3), 26-32 (Hl), 53-55 (H2), and 96-101 (H3) (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987));

[0075] (b) CDRs occurring at amino acid residues 24-34 (LI), 50-56 (L2), 89-97 (L3), 31 -35b (Hl), 50-65 (H2), and 95-102 (H3) (Kabat etal., Sequences of Proteins of Immunological Interest, 5th Ed. Public Health Service, National Institutes of Health, Bethesda, MD (1991)); and

[0076] (c) antigen contacts occurring at amino acid residues 27c-36 (LI), 46-55 (L2), 89-96 (L3), 30-35b (Hl), 47-58 (H2), and 93-101 (H3) (MacCallum et al. J. Mol. Biol. 262: 732-745 (1996)).

[0077] Unless otherwise indicated, the CDRs are determined according to Kabat et al., supra. One of skill in the art will understand that the CDR designations can also be determined according to Chothia, supra, McCallum, supra, or any other scientifically accepted nomenclature system. The numbering of amino acid residues of the VH and VL of the antibody of the invention is made according to Kabat nomenclature if not specifically mentioned otherwise. A skilled person can convert the numbering according to Kabat into other nomenclatures such as Clothia, McCallum, etc.

[0078] “Framework” or “FR” refers to variable domain residues other than complementary determining regions (CDRs). The FR of a variable domain generally consists of four FR domains: FR1, FR2, FR3, and FR4. Accordingly, the CDR and FR sequences generally appear in the following sequence in VH (or VL): FR1-CDR-H1(CDR-L1)-FR2- CDR-H2(CDR-L2)-FR3- CDR-H3(CDR-L3)-FR4.

[0079] In the context of the invention, it is referred to variants of sequences (in particular CDRs). These variants typically comprise one or more amino acid substitutions. It is evident that the variant CDRs are functional variants, i.e. having amino acid sequences that may differ from the reference amino acid sequence but which differing sequence exhibits or maintains the same functional activity as the reference sequence in the context of the described heavy and / or light chain variable domain. Specifically, as used herein, the term same functional activity means that the antibody or antibody binding fragment of the invention comprising one or more variant CDRs will maintain the extraordinary high association rate constant kaand / or the affinity with respect to the binding to HbAlc, the specificity (in particular with respect to the discrimination of HbAlc from structurally related compounds) and / or the coefficient of variation in an immunoassay are maintained.

[0080] The term “epitope” denotes the site on an antigen, either proteinaceous or non-proteinaceous, to which an antibody binds. Epitopes can be formed both from contiguous amino acid stretches (linear epitope) or comprise non-contiguous amino acids (conformational epitope), e.g., coming in spatial proximity due to the folding of the antigen, i.e. by the tertiary folding of a proteinaceous antigen. An epitope comprises at least 3, at least 4, at least 5, at least 6, at least 7, or 8-10 amino acids in a unique spatial conformation.

[0081] “Immunoassays” as used herein are well-established bioanalytical methods in which detection or quantitation of an analyte depends on the reaction of the analyte and at least one analyte-specific binding agent, thus forming an analyte:binding agent complex. In the context of the present invention, at least one of the at least one analyte specific binding agent is an antibody of the invention.

[0082] A “homogeneous immunoassay” is an assay in which the reaction takes place in a single-phase system. A homogeneous immunoassay typically lacks a separation step to distinguish between bound and unbound analyte, i.e. antigen. Typically, the signal is generated by an enzymatic reaction or by a fluorophore attached to the antibody.

[0083] A “turbidimetric inhibition immunoassay” is an assay in which changes in turbidity resulting from analyte-antibody interactions are measured to determine the amount or concentration of the analyte. In this assay, a known quantity of an antigen is mixed with its specific antibody to form an antigen-antibody complex. The test sample containing the analyte of interest - which is typically an antigen similar to the known antigen - is then added into the mixture. The analyte competes with the known antigen for binding sites on the antibody, inhibiting the formation of immune complexes. The degree of turbidity, which is inversely proportional to the concentration of the analyte in the test sample, is measured using a spectrophotometer or similar optical device. By comparing the turbidity of the test sample to that of a standard calibration curve, the concentration of the analyte can be determined.

[0084] A “sample” as used in the context of the present disclosure may be a liquid sample comprising or expected to comprise HbAlc. The sample may in particular be a body fluid, such as, but not restricted to a blood sample, saliva or urine. In embodiments, the sample is a blood sample, such as whole blood, serum or plasma. In embodiments, the sample is serum or plasma. As used herein “affinity” refers to the strength of the sum total of noncovalent interactions between a single binding site of a molecule (e.g., an antibody) and its binding partner (e.g., an antigen). Unless indicated otherwise, as used herein, “binding affinity” refers to intrinsic binding affinity which reflects a 1:1 interaction between members of a binding pair (e.g., antibody and antigen). The affinity of a molecule X for its partner Y can generally be represented by the dissociation equilibrium constant KD). Affinity can be measured by common methods known in the art, including those described herein. Specific illustrative and exemplary methods for measuring binding affinity are described in the following.

[0085] An antibody is said to “bind”, also said to “specifically bind”, to an antigen (e.g. HbAlc) when the antibody has a KD of 100 nM or lower In the following detailed description of the invention, a number of individual elements, characterizing features, techniques and / or steps are disclosed. It is readily recognized that each of these has benefit not only individually when considered or used alone, but also when considered and used in combination with one another. Accordingly, to avoid exceedingly repetitious and redundant passages, this description has refrained from reiterating every possible combination and permutation. Nevertheless, whether expressly recited or not, it is understood that such combinations are entirely within the scope of the presently disclosed subject matter.

[0086] All technical and scientific terms used herein, unless otherwise defined, are intended to have the same meaning as commonly understood by one of ordinary skill in the art. Reference to techniques employed herein are intended to refer to the techniques as commonly understood in the art, including variations on those techniques or substitutions of equivalent techniques that would be apparent to one of skill in the art.

[0087] All amino acid sequences provided herein are presented starting with the most N-terminal residue and ending with the most C-terminal residue, as customarily done in the art, and the one-letter or three-letter code abbreviations as used to identify amino acids throughout the present invention correspond to those commonly used for amino acids.

[0088] In this specification, a number of documents including patent applications and manufacturer’s manuals are cited. The disclosure of these documents, while not considered relevant for the patentability of this invention, is herewith incorporated by reference in its entirety. More specifically, all referenced documents are incorporated by reference to the same extent as if each individual document was specifically and individually indicated to be incorporated by reference.

[0089] In a first aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof.

[0090] It could be shown in various Experiments described herein, including crystallization of the antibody or antigen binding fragment thereof and HbAl c, and calculation of free binding energy between HbAlc and the antibody or antigen binding fragment thereof of the present invention using computer simulations, that the paratope of the antibody or antigen binding fragment thereof binding to HbAlc is defined by CDRs H2, H3, LI and L3. HbAlc itself is a smaller epitope and the interaction between the antibodies or antigen binding fragments thereof of the present invention and HbAlc is mainly defined by CDR-H3, which is a larger CDR-H3 than usual, and the sugar moiety, while the other CDRs contribute less or nothing (CDR-H1 / CDR-L2) to the binding.

[0091] In embodiments, the variant of SEQ ID NO: 2 is having at least 85 %, 90 % or 95 % sequence identity.

[0092] In embodiments, the variant of SEQ ID NO: 3 is having at least 85 %, 90 % or 95 % sequence identity.

[0093] In embodiments, the variant of SEQ ID NO: 4 is having at least 85 %, 90 % or 95 % sequence identity.

[0094] In embodiments, the variant of SEQ ID NO: 6 is having at least 85 %, 90 % or 95 % sequence identity.

[0095] In embodiments, the variant of SEQ ID NO: 2 has

[0096] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19, and / or

[0097] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T. In specific embodiments, the variant of SEQ ID NO: 2 has

[0098] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0099] As shown in the Experiments described herein, some of the amino acids of the paratope can be substituted, in some embodiments even more than one amino acid can be substituted. The calculations of the free binding energy shows that the binding to the epitope depends on only a few amino acids of the paratope, mainly of CDR-H3. Some amino acids of the CD Rs can be substituted by any amino acid without increasing the free binding energy by more than 1.75 kJ / mol and can thus be freely exchanged. Other amino acids, more important for binding, can be only replaced by a more limited number of amino acids without increasing the free binding energy too much, in particular by 3.5 kJ / mol or more. The number of amino acid substitutions of each CDR is mainly limited by the possibility to adopt the conformation needed for binding to HbAlc. More amino acid substitutions might alter the conformation such that a binding is not possible anymore.

[0100] In embodiments, the variant of SEQ ID NO: 3 has

[0101] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0102] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0103] In specific embodiments, the variant of SEQ ID NO: 3 has

[0104] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0105] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0106] In embodiments, the variant of SEQ ID NO: 4 has (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0107] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 4 has

[0108] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0109] In embodiments, the variant of SEQ ID NO: 6 has

[0110] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0111] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0112] In specific embodiments, the variant of SEQ ID NO: 6 has

[0113] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0114] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0115] In specific embodiments, the variant of SEQ ID NO: 6 has

[0116] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 6 has

[0117] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0118] In embodiments, the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof with one amino acid substitution. In particular, the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 2, a CDR-H3 as shown in SEQ ID NO: 3, a CDR-L1 as shown in SEQ ID NO: 4, and a CDR-L3 as shown in SEQ ID NO: 6.

[0119] In embodiments, the variant of the sequence comprised in the antibody or antigen binding fragment thereof is a variant with at most four, in particular at most three, in particular at most two amino acid substitutions, in particular one amino acid substitution.

[0120] In embodiments, the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the sugar moiety and the amino acid residues Vail, His2 and Thr4 as shown in SEQ ID NO: 33 or a variant thereof.

[0121] In embodiments, the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the sugar moiety and the amino acid residues Vail, His2 and Thr4 as shown in SEQ ID NO: 33.

[0122] In embodiments, the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions. In specific embodiments, the variant is a variant with one amino acid substitution.

[0123] In specific embodiments, the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33.

[0124] In embodiments, the antibody or antigen binding fragment thereof binds to an antigen comprising the sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions. In specific embodiments, the variant is a variant with one amino acid substitution.

[0125] In specific embodiments, the antibody or antigen binding fragment thereof binds to an antigen comprising the sequence as shown in SEQ ID NO: 33.

[0126] In embodiments, the variant of SEQ ID NO: 33 is a variant wherein the one or two, in particular one, of the amino acid substitutions is one of the following amino acid substitutions L3V, L3P, T4P or T4N. In specific embodiments, the variant of SEQ ID NO: 33 does not comprise further amino acid substitutions than the one or two amino acid substitutions selected from L3V, L3P, T4P or T4N. In embodiments, all amino acid substitutions of the antibody or antigen binding fragment thereof are conservative amino acid substitutions.

[0127] In embodiments, each of said conservative amino acid substitutions is the substitution of an amino acid with another amino acid selected from its same group, wherein the groups of amino acids are

[0128] a) the nonpolar, hydrophobic amino acids consisting of Gly, Ala, Val, Leu, Ile, Phe, Try, Trp, and Met;

[0129] b) the polar, neutral amino acids consisting of Ser, Thr, Asn, and Gln;

[0130] c) the positively charged, basic amino acids consisting of Arg, Lys, and His, and

[0131] d) the negatively charged, amino acids consisting of Asp and Glu;

[0132] wherein if Cys is to be conservatively substituted, it is substituted with Ser or Ala, and

[0133] wherein if Pro is to be conservatively substituted it is substituted with Ala.

[0134] In embodiments, each of said conservative amino acid substitutions is a highly conservative amino acid substitution selected from

[0135] a) substitution of Ala with Val, Leu, Ile or Gly;

[0136] b) substitution of Arg with Lys;

[0137] c) substitution of Asn with Gin;

[0138] d) substitution of Asp with Glu;

[0139] e) substitution of Cys with Ser;

[0140] f) substitution of Gin with Asn;

[0141] g) substitution of Glu with Asp;

[0142] h) substitution of Gly with Ala; i) substitution of His with Arg;

[0143] j) substitution of Ile with Leu, Val or Ala;

[0144] k) substitution of Leu with Ile, Val or Ala;

[0145] l) substitution of Lys with Arg;

[0146] m) substitution of Met with Leu, Ile or Val;

[0147] n) substitution of Phe with Try or Trp;

[0148] o) substitution of Pro with Ala;

[0149] p) substitution of Ser with Thr;

[0150] q) substitution of Thr with Ser;

[0151] r) substitution of Trp with Phe or Tyr;

[0152] s) substitution of Tyr with Phe or Trp; and

[0153] t) substitution of Val with Leu, Ile or Ala.

[0154] In embodiments, the antibody is a monoclonal antibody.

[0155] In embodiments, the antigen binding fragment is a Fab-fragment, a F(ab')2-fragment, a Fab'-fragment, a single-chain variable fragment (scFv), di-scFv, single-domain antibody, or a protein comprising (VL-CL)nand (VH-CHl)n, wherein n is an integer of 2 to 12 and wherein each (VL-CL) or each (VH-CH1) is connected to the next (VL-CL) or next (VH-CH1) via a peptide linker. In particular, the antigen binding fragment is a Fab-fragment.

[0156] In embodiments, the antibody or antigen binding fragment thereof is a recombinant antibody or antigen binding fragment thereof.

[0157] As demonstrated by the appended Examples, the monoclonal antibodies of the invention are characterized by a particularly low dissociation equilibrium constant (KD) for the binding to HbAlc. This makes the antibodies of the invention particularly suitable for a high throughput immunoassay and a competitive assay format. In embodiments, the antibody or antigen binding fragment thereof binds to HbAlc with a dissociation equilibrium constant, KD, of 20 nM or lower, preferably wherein the KD is measured at 37 °C. In particular, the antibody or antigen binding fragment thereof binds to HbAlc with a dissociation equilibrium constant, KD, of 0.3 nM to 20 nM, preferably wherein the KDis measured at 37 °C.

[0158] Methods for determining the KD of an antibody are known in the art. These values are preferably determined using surface plasmon resonance spectroscopy (SPR). The experimental conditions are preferably as described herein.

[0159] In embodiments, the KD may be determined using kinetic measurements determining the association and dissociation rate constants. Accordingly, the methods may in embodiments be surface plasmon resonance methods.

[0160] Surface plasmon resonance measurements for determining the KD may be conducted at a temperature of 37 °C. The system buffer may be a HBS buffer with a pH 6.2 containing 10 mM HEPES, 150 mM NaCl, 3 mM EDTA, 0.05 % (w / v) Polysorbate 20, in particular when biotinylated screening reagents are used. As sample buffer the latter system buffer supplemented with 1 mg / mL Carboxymethyldextran (CMD) may be used.

[0161] The association rate constant constant ka[M’1], the dissociation rate constant kd[s-1] and the dissociation equilibrium constant KD [M] may be calculated according to a Langmuir 1: 1 model.

[0162] In preferred embodiments, the KD may be determined using surface plasmon resonance spectroscopy (e.g. BIAcore®) and using a direct binding measurement principle, where one of the interactants is immobilized to the sensor surface, the other (the analyte) is free in solution and passed over the surface.

[0163] In an embodiment, the antibody or antigen binding fragment thereof for which the dissociation equilibrium constant is to be determined, (first antibody) is immobilized onto the sensor chip surface via an antibody capture system using a second antibody to bind the first antibody, preferably without interfering with its binding to its antigen. The HbAlc antigen may be biotinylated and pre-incubated with streptavidin, to increase its mass and thus to increase the signal, in particular when only a fragment of HbAlc is used as a screening reagent, i.e. analyte. In another embodiment, the antigen is immobilized onto the sensor chip surface via amine coupling with, for example, EDC / NHS -chemistry. The antibody or antigen binding fragment thereof for which the dissociation equilibrium constant is to be determined is used as a screening reagent, i.e. analyte.

[0164] In embodiments, a GE Healthcare BIAcore™ 8K+, a BIAcore™ B4000 or a BIAcore™ T200 are used for determining the association rate constant ka[M’1], the dissociation rate constant Ay [s’J] and / or the dissociation equilibrium constant KD [M], In a preferred embodiment, the analysis may be conducted with a GE Healthcare Biacore™ 8K instrument and the analysis may be conducted automatically with the Evaluation Insight Software V3.011.15423.

[0165] In embodiments, the antibody or antigen binding fragment thereof binds to HbAlc with an association rate constant (Aa) of 25000 M^s’1or higher. In particular, the antibody or antigen binding fragment thereof binds to HbAlc with an association rate constant (Aa) of 25000 M^s’1to 100000 M’1. These values are preferably determined using surface plasmon resonance spectroscopy. The experimental conditions are preferably as described herein, in particular the same as described for the KD.

[0166] In embodiments, the antibody or antigen binding fragment thereof binds to HbAlc with a dissociation rate constant kd of 0.00024 s’1or lower. In particular, the antibody or antigen binding fragment thereof binds to HbAlc with a dissociation rate constant (Ay) of 0.00024 s’1to 0.001 s’1. These values are preferably determined using surface plasmon resonance spectroscopy. The experimental conditions are preferably as described herein, in particular the same as described for the KD.

[0167] In embodiments, the antibody or antigen binding fragment thereof binds to HbAlc with a t / 2 diss of 12 min or more. In particular, the antibody or antigen binding fragment thereof binds to HbAl c with a t / 2 diss of 12 min to 46 min. These values are preferably determined using surface plasmon resonance spectroscopy. The experimental conditions are preferably as described herein, in particular the same as described for the KD.

[0168] The inventors have surprisingly found that using the presently claimed antibodies or antigen binding fragments thereof alone without other antibodies leads to a particularly low coefficient of variation in immunoassays, in particular turbidimetric inhibition immunoassays, in comparison to other antibodies or antigen binding fragments used alone, or lead to a similar coefficient of variation when compared to polyclonal antibodies. When the antibody or antigen binding fragment thereof may be used in a turbidimetric inhibition immunoassay using a sample comprising a normal physiological concentration of HbAlc, the immunoassay may have a coefficient of variation of 1.5 % or lower. In particular, when the antibody or antigen binding fragment thereof may be used in a turbidimetric inhibition immunoassay using a sample comprising a normal physiological concentration of HbAlc, the immunoassay may have a coefficient of variation of 0.8 % or lower. The sample may be comprising HbAlc at a concentration lower than 6.0 %, wherein the percentage is the amount of HbAlc compared to the total amount of hemoglobin as measured by turbidimetric inhibition immunoassay. According to a report of the World Health Organization (WHO) (Use of Glycated Hemoglobin (HbAlc) in the Diagnosis of Diabetes Mellitus, 2011, WHO) HbAlc levels equal or above 6.0 % are associated with higher risk of diabetes and might require intervention, and are therefore considered to be abnormal, while levels lower are considered a normal physiological concentration of HbAlc.

[0169] When the antibody or antigen binding fragment thereof may be used in a turbidimetric inhibition immunoassay using a sample comprising an abnormal physiological concentration of HbAlc, the immunoassay may have a coefficient of variation of 1.5 % or lower, in particular 1.2 % or lower. The sample may be comprising HbAlc at a concentration higher than or equal to 6.0 %, wherein the percentage is the amount of HbAl c compared to the total amount of hemoglobin as measured by turbidimetric inhibition immunoassay.

[0170] In embodiments, the antibody or antigen binding fragment thereof binds to glycated hemoglobin subunit beta.

[0171] In embodiments, the antibody or antigen binding fragment thereof binds to an antigen comprising or consisting of SEQ ID NO: 41 or a variant thereof with one amino acid substitution. In particular, the variant may be L3 V, L3P, T4P or T4N.

[0172] In embodiments, the antibody or antigen binding fragment thereof does not bind to non-glycated hemoglobin subunit beta. In particular, the antibody or antigen binding fragment thereof does not bind to hemoglobin A0 (HbAO). In particular, the antibody or antigen binding fragment thereof does not bind to hemoglobin A0 (HbAO) when tested with ELISA, in particular binding HbAO with an OD of less than 0.1, wherein HbAO is immobilized in a well of a standard 96-well plate with 100 µl of 31.25 ng / ml HbAO and to which well, after removal of the previous solution, are 30 µl of B-cell culture supernatant added. In another embodiment, the antibody or antigen binding fragment thereof does not bind to hemoglobin AO (HbAO), when the affinity to HbAO is higher than 100 nM. The affinity may be measured similarly as the KD, ka, kd or affinity of binding to HbAlc is measured, as described in this application.

[0173] In a second aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof.

[0174] All embodiments mentioned for the first aspect of the invention apply for the second aspect of the invention and vice versa.

[0175] In embodiments, the variant of SEQ ID NO: 8 is having at least 85 %, 90 % or 95 % sequence identity.

[0176] In embodiments, the variant of SEQ ID NO: 9 is having at least 85 %, 90 % or 95 % sequence identity.

[0177] In embodiments, the variant of SEQ ID NO: 10 is having at least 85 %, 90 % or 95 % sequence identity.

[0178] In embodiments, the variant of SEQ ID NO: 12 is having at least 85 %, 90 % or 95 % sequence identity.

[0179] In embodiments, the variant of SEQ ID NO: 8 has

[0180] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19;

[0181] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T; and / or (iii) deletion of at most three amino acids. In specific embodiments, the variant of SEQ ID NO: 8 has

[0182] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0183] In embodiments, the variant of SEQ ID NO: 9 has

[0184] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0185] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0186] In specific embodiments, the variant of SEQ ID NO: 9 has

[0187] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0188] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0189] In embodiments, the variant of SEQ ID NO: 10 has

[0190] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 10 has

[0191] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0192] In embodiments, the variant of SEQ ID NO: 12 has

[0193] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0194] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0195] In specific embodiments, the variant of SEQ ID NO: 12 has

[0196] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0197] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In specific embodiments, the variant of SEQ ID NO: 12 has

[0198] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 12 has

[0199] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0200] In embodiments, the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof with one amino acid substitution. In particular, the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 8, a CDR-H3 as shown in SEQ ID NO: 9, a CDR-L1 as shown in SEQ ID NO: 10, and a CDR-L3 as shown in SEQ ID NO: 12. In a third aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof.

[0201] All embodiments mentioned for the first and / or second aspect of the invention apply for the third aspect of the invention and vice versa.

[0202] In embodiments, the variant of SEQ ID NO: 14 is having at least 85 %, 90 % or 95 % sequence identity.

[0203] In embodiments, the variant of SEQ ID NO: 15 is having at least 85 %, 90 % or 95 % sequence identity.

[0204] In embodiments, the variant of SEQ ID NO: 16 is having at least 85 %, 90 % or 95 % sequence identity.

[0205] In embodiments, the variant of SEQ ID NO: 18 is having at least 85 %, 90 % or 95 % sequence identity.

[0206] In embodiments, the variant of SEQ ID NO: 14 has

[0207] (i) at most four, in particular two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 16;

[0208] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0209] In specific embodiments, the variant of SEQ ID NO: 14 has

[0210] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 16.

[0211] In embodiments, the variant of SEQ ID NO: 15 has

[0212] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0213] In specific embodiments, the variant of SEQ ID NO: 15 has

[0214] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0215] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0216] In embodiments, the variant of SEQ ID NO: 16 has

[0217] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0218] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 16 has

[0219] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0220] In embodiments, the variant of SEQ ID NO: 18 has (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0221] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0222] In specific embodiments, the variant of SEQ ID NO: 18 has

[0223] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0224] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In particular embodiments, the variant of SEQ ID NO: 18 has

[0225] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular embodiments, the variant of SEQ ID NO: 18 has (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0226] In embodiments, the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof with one amino acid substitution. In particular, the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 14, a CDR-H3 as shown in SEQ ID NO: 15, a CDR-L1 as shown in SEQ ID NO: 16 and a CDR-L3 as shown in SEQ ID NO: 18.

[0227] In a fourth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 1 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 5 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof.

[0228] All embodiments mentioned for the first, second and / or third aspect of the invention apply for the fourth aspect of the invention and vice versa.

[0229] In embodiments, the variant of SEQ ID NO: 1 is having at least 85 %, 90 % or 95 % sequence identity.

[0230] In embodiments, the variant of SEQ ID NO: 2 is having at least 85 %, 90 % or 95 % sequence identity.

[0231] In embodiments, the variant of SEQ ID NO: 3 is having at least 85 %, 90 % or 95 % sequence identity.

[0232] In embodiments, the variant of SEQ ID NO: 4 is having at least 85 %, 90 % or 95 % sequence identity.

[0233] In embodiments, the variant of SEQ ID NO: 5 is having at least 85 %, 90 % or 95 % sequence identity.

[0234] In embodiments, the variant of SEQ ID NO: 6 is having at least 85 %, 90 % or 95 % sequence identity.

[0235] In embodiments, the variant of SEQ ID NO: 1 has

[0236] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0237] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0238] In specific embodiments, the variant of SEQ ID NO: 1 has

[0239] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0240] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0241] In embodiments, the variant of SEQ ID NO: 2 has (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19, and / or

[0242] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T. In specific embodiments, the variant of SEQ ID NO: 2 has

[0243] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0244] In embodiments, the variant of SEQ ID NO: 3 has

[0245] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0246] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0247] In specific embodiments, the variant of SEQ ID NO: 3 has

[0248] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0249] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0250] In embodiments, the variant of SEQ ID NO: 4 has

[0251] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0252] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 4 has

[0253] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0254] In embodiments, the variant of SEQ ID NO: 6 has

[0255] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0256] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0257] In specific embodiments, the variant of SEQ ID NO: 6 has

[0258] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0259] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In specific embodiments, the variant of SEQ ID NO: 6 has

[0260] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 6 has

[0261] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0262] In embodiments, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 1 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 5 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof with one amino acid substitution. In particular, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 1, a CDR-H2 as shown in SEQ ID NO: 2, a CDR-H3 as shown in SEQ ID NO: 3, a CDR-L1 as shown in SEQ ID NO: 4, a CDR-L2 as shown in SEQ ID NO: 5 and a CDR-L3 as shown in SEQ ID NO: 6.

[0263] In a fifth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 7 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 11 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof.

[0264] All embodiments mentioned for the first, second, third and / or fourth aspect of the invention apply for the fifth aspect of the invention and vice versa.

[0265] In embodiments, the variant of SEQ ID NO: 7 is having at least 85 %, 90 % or 95 % sequence identity.

[0266] In embodiments, the variant of SEQ ID NO: 8 is having at least 85 %, 90 % or 95 % sequence identity.

[0267] In embodiments, the variant of SEQ ID NO: 9 is having at least 85 %, 90 % or 95 % sequence identity.

[0268] In embodiments, the variant of SEQ ID NO: 10 is having at least 85 %, 90 % or 95 % sequence identity.

[0269] In embodiments, the variant of SEQ ID NO: 11 is having at least 85 %, 90 % or 95 % sequence identity.

[0270] In embodiments, the variant of SEQ ID NO: 12 is having at least 85 %, 90 % or 95 % sequence identity. In embodiments, the variant of SEQ ID NO: 7 has

[0271] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0272] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0273] In specific embodiments, the variant of SEQ ID NO: 7 has

[0274] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0275] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0276] In embodiments, the variant of SEQ ID NO: 8 has

[0277] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19;

[0278] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T; and / or

[0279] (iii) deletion of at most three amino acids. In specific embodiments, the variant of SEQ ID NO: 8 has

[0280] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0281] In embodiments, the variant of SEQ ID NO: 9 has

[0282] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0283] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0284] In specific embodiments, the variant of SEQ ID NO: 9 has

[0285] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0286] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0287] In embodiments, the variant of SEQ ID NO: 10 has

[0288] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0289] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 10 has

[0290] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0291] In embodiments, the variant of SEQ ID NO: 12 has

[0292] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0293] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W. In specific embodiments, the variant of SEQ ID NO: 12 has

[0294] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0295] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In specifc embodiments, the variant of SEQ ID NO: 12 has

[0296] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 12 has

[0297] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0298] In embodiments, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 7 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 11 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof with one amino acid substitution. In particular, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 7, a CDR-H2 as shown in SEQ ID NO: 8, a CDR-H3 as shown in SEQ ID NO: 9, a CDR-L1 as shown in SEQ ID NO: 10, a CDR-L2 as shown in SEQ ID NO: 11 and a CDR-L3 as shown in SEQ ID NO: 12.

[0299] In a sixth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 13 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 17 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof.

[0300] All embodiments mentioned for the first, second, third, fourth and / or fifth aspect of the invention apply for the sixth aspect of the invention and vice versa. In embodiments, the variant of SEQ ID NO: 13 is having at least 85 %, 90 % or 95 % sequence identity.

[0301] In embodiments, the variant of SEQ ID NO: 14 is having at least 85 %, 90 % or 95 % sequence identity.

[0302] In embodiments, the variant of SEQ ID NO: 15 is having at least 85 %, 90 % or 95 % sequence identity.

[0303] In embodiments, the variant of SEQ ID NO: 16 is having at least 85 %, 90 % or 95 % sequence identity.

[0304] In embodiments, the variant of SEQ ID NO: 17 is having at least 85 %, 90 % or 95 % sequence identity.

[0305] In embodiments, the variant of SEQ ID NO: 18 is having at least 85 %, 90 % or 95 % sequence identity.

[0306] In embodiments, the variant of SEQ ID NO: 13 has

[0307] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0308] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0309] In specific embodiments, the variant of SEQ ID NO: 13 has

[0310] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0311] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0312] In embodiments, the variant of SEQ ID NO: 14 has

[0313] (i) at most four, in particular two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 16;

[0314] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0315] In specific embodiments, the variant of SEQ ID NO: 14 has

[0316] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 16.

[0317] In embodiments, the variant of SEQ ID NO: 15 has

[0318] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0319] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0320] In specific embodiments, the variant of SEQ ID NO: 15 has

[0321] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0322] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0323] In embodiments, the variant of SEQ ID NO: 16 has (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0324] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 16 has

[0325] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0326] In embodiments, the variant of SEQ ID NO: 18 has

[0327] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0328] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0329] In specific embodiments, the variant of SEQ ID NO: 18 has

[0330] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0331] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In specific embodiments, the variant of SEQ ID NO: 18 has

[0332] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 18 has

[0333] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0334] In embodiments, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 13 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 17 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof with one amino acid substitution. In particular, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 13, a CDR-H2 as shown in SEQ ID NO: 14, a CDR-H3 as shown in SEQ ID NO: 15, a CDR-L1 as shown in SEQ ID NO: 16, a CDR-L2 as shown in SEQ ID NO: 17 and a CDR-L3 as shown in SEQ ID NO: 18.

[0335] In a seventh aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 19 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 20 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 21 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 22 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 23 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 24 or variant thereof.

[0336] All embodiments mentioned for the first to fifth and / or sixth aspect of the invention apply for the seventh aspect of the invention and vice versa.

[0337] In embodiments, the variant of SEQ ID NO: 19 is having at least 85 %, 90 % or 95 % sequence identity.

[0338] In embodiments, the variant of SEQ ID NO: 20 is having at least 85 %, 90 % or 95 % sequence identity.

[0339] In embodiments, the variant of SEQ ID NO: 21 is having at least 85 %, 90 % or 95 % sequence identity.

[0340] In embodiments, the variant of SEQ ID NO: 22 is having at least 85 %, 90 % or 95 % sequence identity.

[0341] In embodiments, the variant of SEQ ID NO: 23 is having at least 85 %, 90 % or 95 % sequence identity.

[0342] In embodiments, the variant of SEQ ID NO: 24 is having at least 85 %, 90 % or 95 % sequence identity.

[0343] In embodiments, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 19 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 20 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 21 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 22 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 23 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 24 or variant thereof with one amino acid substitution. In particular, the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 19, a CDR-H2 as shown in SEQ ID NO: 20, a CDR-H3 as shown in SEQ ID NO: 21, a CDR-L1 as shown in SEQ ID NO: 22, a CDR-L2 as shown in SEQ ID NO: 23 and a CDR-L3 as shown in SEQ ID NO: 24.

[0344] In an eighth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: 25 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 26 or variant thereof.

[0345] All embodiments mentioned for the first to sixth and / or seventh aspect of the invention apply for the eighth aspect of the invention and vice versa.

[0346] In embodiments, the variant of SEQ ID NO: 25 is having at least 85 %, 90 % or 95 % sequence identity.

[0347] In embodiments, the variant of SEQ ID NO: 26 is having at least 85 %, 90 % or 95 % sequence identity.

[0348] In embodiments, the heavy chain variable domain comprises

[0349] a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 1 or a variant thereof,

[0350] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 2 or a variant thereof, and

[0351] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 3 or a variant thereof; and / or

[0352] the light chain variable domain comprises

[0353] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 4 or a variant thereof, a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 5 or a variant thereof, and

[0354] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 6 or a variant thereof.

[0355] In embodiments, the variant of SEQ ID NO: 1 has

[0356] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0357] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0358] In specific embodiments, the variant of SEQ ID NO: 1 has

[0359] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0360] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0361] In embodiments, the variant of SEQ ID NO: 2 has

[0362] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19, and / or

[0363] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T. In specific embodiments, the variant of SEQ ID NO: 2 has

[0364] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0365] In embodiments, the variant of SEQ ID NO: 3 has

[0366] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0367] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0368] In specific embodiments, the variant of SEQ ID NO: 3 has

[0369] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0370] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0371] In embodiments, the variant of SEQ ID NO: 4 has

[0372] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0373] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 4 has

[0374] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0375] In embodiments, the variant of SEQ ID NO: 6 has

[0376] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0377] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W. In specific embodiments, the variant of SEQ ID NO: 6 has

[0378] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0379] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In a particular embodiment, the variant of SEQ ID NO: 6 has

[0380] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 6 has

[0381] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0382] In a ninth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: SEQ ID NO: 27 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 28 or variant thereof.

[0383] All embodiments mentioned for the first to seventh and / or eighth aspect of the invention apply for the ninth aspect of the invention and vice versa.

[0384] In embodiments, the variant of SEQ ID NO: 27 is having at least 85 %, 90 % or 95 % sequence identity.

[0385] In embodiments, the variant of SEQ ID NO: 28 is having at least 85 %, 90 % or 95 % sequence identity.

[0386] In embodiments, the heavy chain variable domain comprises

[0387] a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 7 or a variant thereof,

[0388] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 8 or a variant thereof, and

[0389] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 9 or a variant thereof; and / or the light chain variable domain comprises

[0390] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 10 or a variant thereof,

[0391] a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 11 or a variant thereof, and

[0392] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 12 or a variant thereof.

[0393] In embodiments, the variant of SEQ ID NO: 7 has

[0394] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0395] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0396] In specific embodiments, the variant of SEQ ID NO: 7 has

[0397] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0398] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0399] In embodiments, the variant of SEQ ID NO: 8 has

[0400] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19;

[0401] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T; and / or

[0402] (iii) deletion of at most three amino acids. In specific embodiments, the variant of SEQ ID NO: 8 has

[0403] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0404] In embodiments, the variant of SEQ ID NO: 9 has (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0405] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0406] In specific embodiments, the variant of SEQ ID NO: 9 has

[0407] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0408] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0409] In embodiments, the variant of SEQ ID NO: 10 has

[0410] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0411] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 10 has

[0412] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10. In embodiments, the variant of SEQ ID NO: 12 has

[0413] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0414] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0415] In specific embodiments, the variant of SEQ ID NO: 12 has

[0416] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0417] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In particular embodiment, the variant of SEQ ID NO: 12 has

[0418] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 12 has

[0419] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0420] In an tenth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: 29 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 30 or variant thereof.

[0421] All embodiments mentioned for the first to eighth and / or ninth aspect of the invention apply for the tenth aspect of the invention and vice versa.

[0422] In embodiments, the variant of SEQ ID NO: 29 is having at least 85 %, 90 % or 95 % sequence identity.

[0423] In embodiments, the variant of SEQ ID NO: 30 is having at least 85 %, 90 % or 95 % sequence identity.

[0424] In embodiments, the heavy chain variable domain comprises a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 13 or a variant thereof,

[0425] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 14 or a variant thereof, and

[0426] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 15 or a variant thereof; and / or

[0427] the light chain variable domain comprises

[0428] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 16 or a variant thereof,

[0429] a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 17 or a variant thereof, and

[0430] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 18 or a variant thereof.

[0431] In embodiments, the variant of SEQ ID NO: 13 has

[0432] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0433] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0434] In specific embodiments, the variant of SEQ ID NO: 13 has

[0435] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0436] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0437] In embodiments, the variant of SEQ ID NO: 14 has

[0438] (i) at most four, in particular two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 16;

[0439] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0440] In specific embodiments, the variant of SEQ ID NO: 14 has

[0441] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 16.

[0442] In embodiments, the variant of SEQ ID NO: 15 has

[0443] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0444] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0445] In specific embodiments, the variant of SEQ ID NO: 15 has

[0446] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0447] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0448] In embodiments, the variant of SEQ ID NO: 16 has (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0449] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L. In specific embodiments, the variant of SEQ ID NO: 16 has

[0450] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0451] In embodiments, the variant of SEQ ID NO: 18 has

[0452] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0453] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0454] In specific embodiments, the variant of SEQ ID NO: 18 has

[0455] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0456] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. In particular embodiments, the variant of SEQ ID NO: 18 has

[0457] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. In particular, the variant of SEQ ID NO: 18 has

[0458] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0459] In an eleventh embodiment, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin A1c (HbA1c) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: 31 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 32 or variant thereof. All embodiments mentioned for any of the first to ninth and / or tenth aspect of the invention apply for the eleventh aspect of the invention and vice versa.

[0460] In embodiments, the variant of SEQ ID NO: 31 is having at least 85 %, 90 % or 95 % sequence identity.

[0461] In embodiments, the variant of SEQ ID NO: 32 is having at least 85 %, 90 % or 95 % sequence identity.

[0462] In embodiments, the heavy chain variable domain comprises

[0463] a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 19 or a variant thereof,

[0464] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 20 or a variant thereof, and

[0465] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 21 or a variant thereof; and / or

[0466] the light chain variable domain comprises

[0467] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 22 or a variant thereof,

[0468] a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 23 or a variant thereof, and

[0469] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 24 or a variant thereof.

[0470] In a twelfth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc), wherein the antibody or antigen binding fragment thereof binds to an epitope comprising the sugar moiety, the amino acid residues Vail, His2 and Thr4 as shown in SEQ ID NO: 33 or a variant thereof.

[0471] All embodiments mentioned for any of the first to tenth and / or eleventh aspect of the invention apply for the twelfth aspect of the invention and vice versa.

[0472] In embodiments, the variant of SEQ ID NO: 33 is a variant with one amino acid substitution. In embodiments, the antibody or antigen binding fragment thereof binds to an epitope comprising the sugar moiety, the amino acid residues Vail, His2 and Thr4 as shown in SEQ ID NO: 33.

[0473] In a thirteenth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc), wherein the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions.

[0474] All embodiments mentioned for any of the first to eleventh and / or twelfth aspect of the invention apply for the thirteenth aspect of the invention and vice versa.

[0475] In embodiments, the variant of SEQ ID NO: 33 is a variant with one amino acid substitution. In particular, the substitution is a conservative amino acid substitution.

[0476] In embodiments, the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33.

[0477] In a fourteenth aspect, the present invention relates to an antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc), wherein the antibody or antigen binding fragment thereof binds to an antigen comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions.

[0478] All embodiments mentioned for any of the first to twelfth and / or thirteenth aspect of the invention apply for the fourteenth aspect of the invention and vice versa.

[0479] In embodiments, the variant is a variant with one amino acid substitution.

[0480] In embodiments, the antibody or antigen binding fragment thereof binds to an antigen comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33.

[0481] In a fifteenth aspect, the present invention relates to a nucleic acid coding for at least a part of an antibody or antigen binding fragment thereof of the present invention.

[0482] All embodiments mentioned for any of the first to thirteenth and / or fourteenth aspect of the invention apply for the fifteenth aspect of the invention and vice versa.

[0483] In embodiments, the nucleic acid is DNA or RNA. In particular, the nucleic acid is DNA. In embodiments, the part of an antibody or antigen binding fragment thereof is at least one CDR, at least one light chain variable domain or at least one heavy chain variable domain. In particular the part thereof is at least all three CDRs of the heavy or light chain variable domain. In another embodiment, the part is the heavy or light chain, i.e. including the constant domain.

[0484] In other embodiments, the nucleic acid encodes for all six CDRs of the heavy and light chain of an antibody or antigen binding fragment thereof of the present invention.

[0485] In a sixteenth aspect, the present invention relates to an expression construct comprising a nucleic acid of the present invention.

[0486] All embodiments mentioned for any of the first to fourteenth and / or fifteenth aspect of the invention apply for the sixteenth aspect of the invention and vice versa.

[0487] With regard to the term "expression construct comprising" as used herein, it is understood in the art that further nucleic acid sequences are present in the expression constructs that are necessary and / or sufficient for desired expression in the host cell, e.g. to direct the host cell express the antibody or antigen binding fragment of the invention. Such further nucleic acid sequences include but are not limited to sequences controlling expression of a desired sequence in the particular cell system. For example, the vectors may comprise the nucleic acid molecule encoding an antibody or antibody antigen binding fragment of the invention operably linked and / or under the control of regulatory sequences. The term "regulatory sequence" refers to DNA sequences that are necessary to effect the expression of coding sequences to which they are operably linked. The term “control sequence” is intended to include, at a minimum, all components the presence of which may also be necessary for expression, and may further include additional advantageous components, e.g., to allow replication. As is understood in the art, the nature of such regulatory and control sequences differs depending upon the host organism. For example, in prokaryotes, control sequences generally include promoters, ribosomal binding sites, and terminators. In eukaryotes control sequences generally include promoters, terminators and, in some instances, enhancers, transactivators and / or transcription factors.

[0488] In embodiments, the expression construct comprises a promoter, a terminator, a 5' UTR and / or a 3' UTR. In some embodiments, the promoter is an inducible promoter. In some embodiments, the promoter is a constitutive promoter. In a seventeenth aspect, the present invention relates to a vector comprising an expression construct of the present invention.

[0489] All embodiments mentioned for any of the first to fifteenth and / or sixteenth aspect of the invention apply for the seventeenth aspect of the invention and vice versa.

[0490] As used herein, the term "vector" relates to a circular or linear nucleic acid molecule that can autonomously replicate in a host cell into which it has been introduced. Non-limiting examples of vectors suitable for use in the present invention include cosmids, plasmids (e.g., naked or contained in liposomes), viruses (e.g., lentiviruses, retroviruses, adenoviruses, and adeno-associated viruses) and bacteriophages. However, the art provides many suitable vectors, the choice of which depends on the desired function. The development and use of suitable vectors is well documented in the art; see, for example, the techniques described in Sambrook and Russel “Molecular Cloning, A Laboratory Manual”, Cold Spring Harbor Laboratory, N. Y. (2001) and Ausubel, “Current Protocols in Molecular Biology”, Green Publishing Associates and Wiley Interscience, N. Y. (1989), (1994).

[0491] With regard to the term "vector comprising" as used herein, it is understood in the art that further nucleic acid sequences are present in the vectors that are necessary and / or sufficient for desired vector activity in the host cell, e.g. drive replication of the vector (and, thus the encoding nucleic acid sequences) and / or to direct the host cell express the antibody or antigen binding fragment of the invention. Such further nucleic acid sequences include but are not limited to sequences controlling vector replication in the particular cell system. For example, the vectors may comprise the nucleic acid molecule encoding an antibody or antibody antigen binding fragment of the invention operably linked and / or under the control of regulatory sequences. The term "regulatory sequence" refers to DNA sequences that are necessary to effect the expression of coding sequences to which they are operably linked. The term “control sequence” is intended to include, at a minimum, all components the presence of which may also be necessary for expression, and may further include additional advantageous components, e.g., to allow replication. As is understood in the art, the nature of such regulatory and control sequences differs depending upon the host organism. For example, in prokaryotes, control sequences generally include promoters, ribosomal binding sites, and terminators. In eukaryotes control sequences generally include promoters, terminators and, in some instances, enhancers, transactivators and / or transcription factors. In embodiments, the vector comprises at least one selection marker and / or at least one origin of replication.

[0492] In embodiments, the vector is a viral vector, artificial chromosome or a plasmid.

[0493] The vectors of use in the present invention are preferably expression vectors. An expression vector is capable of directing the replication and the expression of the nucleic acid molecule of the invention in a host cell and, accordingly, provides for the expression of, e.g., the heavy chain and / or light chain variable domains of the monoclonal antibodies specifically binding to T4 as disclosed herein. In some embodiments, the vector may comprise further sequences to ensure that not only the heavy and light chain variable domains are expressed, but also the remaining heavy and light chain constant regions such that a full-length IgG antibody is expressed comprising the heavy and light chain variable domains of the invention. Suitable expression vectors have been widely described in the literature and the determination of the appropriate expression vector for a particular cell system can be readily made by the skilled person using routine methods. Preferably, the vectors disclosed herein comprise a recombinant polynucleotide (i.e., a nucleic acid sequence encoding the monoclonal antibody according to the invention) as well as expression operably linked control sequences. The vectors as provided herein preferably further comprise a promoter. The herein described vectors may also comprise a selection marker gene and a replication-origin ensuring replication in the host Moreover, the herein provided vectors may also comprise a termination signal for transcription. Expression vectors as known in the art may drive transient or constitutive expression in a host cell.

[0494] The nucleic acid molecules and / or vectors of the invention can be designed for transfection into prokaryotic or eukaryotic host cells by any means known in the art or described herein. Nonlimiting examples of suitable methods include chemical based methods (polyethylenimine, calcium phosphate, liposomes, DEAE-dextrane, nucleofection), nonchemical methods (electroporation, sonoporation, optical transfection, gene electrotransfer, hydrodynamic delivery or naturally occurring transformation upon contacting cells with the nucleic acid molecule of the invention), particle-based methods (gene gun, magnetofection, impalefection) phage vectorbased methods and viral methods. For example, expression vectors derived from viruses such as retroviruses, vaccinia virus, adeno-associated virus, herpes viruses, Semliki Forest Virus or bovine papilloma virus, may be used for transfection of the nucleic acid molecules into targeted cell population. Additionally, baculoviral systems can also be used as vector in eukaryotic expression system for the nucleic acid molecules of the invention.

[0495] The term “prokaryote” is meant to include all bacteria which can be transformed, transduced or transfected with DNA or DNA or RNA molecules for the expression of a protein of the invention. Prokaryotic hosts may include gram negative as well as gram positive bacteria such as, for example, E. coli, S. typhimurium, Serratia marcescens, Corynebacterium (glutamicum), Pseudomonas (fluorescens), Lactobacillus, Streptomyces, Salmonella and Bacillus subtilis. The term "eukaryotic" is meant to include yeast, higher plant, insect and mammalian cells. Nonlimiting examples of mammalian host cells typically used in the art include, Hela, HEK293, H9, Per. C6 and Jurkat cells, mouse NIH3T3, NS / 0, SP2 / 0 and C127 cells, COS cells, e.g. COS 1 or COS 7, CV1, quail QC1-3 cells, mouse L cells, mouse sarcoma cells, Bowes melanoma cells and Chinese hamster ovary (CHO) cells.

[0496] In a eighteenth aspect, the present invention relates to a cell comprising an antibody or antigen binding fragment thereof, a nucleic acid, an expression constructor a vector of the present invention.

[0497] All embodiments mentioned for any of the first to sixteenth and / or seventeenth aspect of the invention apply for the eighteenth aspect of the invention and vice versa.

[0498] In embodiments, the cell is a eukaryotic cell, in particular a human cell.

[0499] In a particular embodiment, the cell is a HEK cell. In another particular embodiment the host cell is a CHO cell.

[0500] When vectors comprising expression constructs of the invention as disclosed herein are introduced into host cells, the antibodies or antigen binding fragments are produced by culturing the host cells for a period of time sufficient to allow for expression of the antibody or antigen binding fragment in the host cell or, preferably, to allow for secretion of the antibody or antigen binding fragment into the culture medium in which the host cells are grown. Antibodies and / or antigen binding fragments can be recovered from the culture medium using standard protein purification methods. Methods for purification of antibodies are well known in the art. Accordingly, the invention also provides a method for the production of an antibody or antigen binding fragment thereof binding to HbAlc as disclosed herein. The method comprises culturing a host cell of the invention under suitable conditions and isolating the antibody or antigen binding fragment thereof produced. By purification steps, e.g. as described in the appended Examples an isolated monoclonal antibody of the invention can be derived. The invention further provides an antibody or an antigen binding fragment obtainable by any of the methods disclosed herein.

[0501] The transformed host cells can be grown in bioreactors and cultured according to techniques known in the art to achieve optimal cell growth. The antibody and / or antigen binding fragment of the invention can then be isolated from the cell fraction or growth medium by any conventional means such, but not limited to, affinity chromatography (for example using a fusion-tag such as the Strep-tag II or the His6tag), gel filtration (size exclusion chromatography), anion exchange chromatography, cation exchange chromatography, hydrophobic interaction chromatography, high pressure liquid chromatography (HPLC), reversed phase HPLC or immunoprecipitation.

[0502] It will be appreciated that variations on the above procedures are within the scope of the present invention. For example, recombinant DNA technology may be used to remove or modify the DNA sequences encoding the antibodies and / or antigen binding fragments disclosed herein, e.g. encoding the heavy and / or light chain variable domains as defined herein above. For example, recombinant DNA technology may be used to remove parts of the encoding sequence(s) that are not necessary for maintaining specific and selective binding to the antigen(s) of interest. The molecules expressed from such truncated DNA molecules are also encompassed by the antibodies of the invention. Additionally, also provided are multivalent antibodies comprising a heavy and / or a light chain variable domain of the invention (e.g. forming and antibody Fv domain that binds HbAlc) at least twice (preferably four, five, six, seven or eight times).

[0503] Antibody derivatives can be produced, for example, by adding exogenous sequences to modify immunogenicity or reduce, enhance or modify binding, affinity, association rate constant, dissociation rate constant, avidity, specificity, half-life, or any other suitable characteristic.

[0504] Also provided are humanized versions of the antibodies disclosed herein, i.e. comprising the CDRs of the heavy and or light chains as disclosed herein above. As well known in the art, “humanization” (to produce a humanized version of a parent antibody) refers to recombinant engineering an antibody using CDRs derived from a non-human donor immunoglobulin in the context of human derived framework and constant domains. During the engineering, framework and / or CDR residues may be altered to preserve binding affinity and activity, e.g. specificity for HbAlc. Methods to humanize antibodies are well known in the art, e.g. as disclosed in Queen et al., Proc. Natl. Acad Sci USA 86(1989), 10029-10032; Hodgson et al., Bio / Technology 9(1991) 421.

[0505] In a nineteenth aspect, the present invention relates to a composition comprising an antibody or antigen binding fragment thereof of the present invention and an acceptable carrier.

[0506] All embodiments mentioned for any of the first to seventeenth and / or eighteenth aspect of the invention apply for the nineteenth aspect of the invention and vice versa.

[0507] In embodiments, the acceptable carrier is water or an aqueous buffered solution.

[0508] In preferred embodiments, the composition is for use in an in vitro diagnostic test for detecting HbAlc.

[0509] In embodiments, the composition of the invention is a composition for in vitro detection (preferably quantification) of HbAlc in a sample, preferably using an immunoassay. In embodiments, the immunoassay is a homogeneous immunoassay. In embodiments, the immunoassay is a competitive immunoassay.

[0510] In a twentieth aspect, the present invention relates to a diagnostic composition comprising an antibody or antigen binding fragment thereof of the present invention and an acceptable carrier.

[0511] All embodiments mentioned for any of the first to eighteenth and / or nineteenth aspect of the invention apply for the twentieth aspect of the invention and vice versa.

[0512] In a twenty-first aspect, the present invention relates to a kit comprising an antibody or antigen binding fragment thereof, a composition or a diagnostic composition of the present invention.

[0513] All embodiments mentioned for any of the first to nineteenth and / or twentieth aspect of the invention apply for the twenty-first aspect of the invention and vice versa.

[0514] In embodiments, the kit comprises an antibody or antigen binding fragment thereof, a composition or a diagnostic composition of the present invention in a container. Suitable container for a solid or liquid composition are known to the skilled person and may be of glass, optionally with a plastic cap, or plastic.

[0515] A kit of the present invention may be used for determining and / or quantifying the amount or concentration of HbAlc in a sample.

[0516] In a twenty-second aspect, the present invention relates to a use of an antibody or antigen binding fragment thereof, a composition or a diagnostic composition of the present invention for determining the amount or concentration of HbAlc in a sample.

[0517] All embodiments mentioned for any of the first to twentieth and / or twenty-first aspect of the invention apply for the twenty-second aspect of the invention and vice versa.

[0518] In embodiments, the sample is a biological fluid sample. In particular, the biological fluid sample is whole blood, in particular capillary or venous blood, serum, blood plasma, saliva or interstitial fluid.

[0519] In a twenty-third aspect, the present invention relates to a method of determining the levels of HbAlc in a sample comprising a step of bringing the sample into contact with an antibody or antigen binding fragment thereof of the present invention.

[0520] All embodiments mentioned for any of the first to twenty-first and / or twenty-second aspect of the invention apply for the twenty-third aspect of the invention and vice versa.

[0521] In a twenty-fourth aspect, the present invention relates to a method of determining the ratio of HbAlc in a sample comprising the following steps:

[0522] (a) adding to the sample an antibody or antigen binding fragment thereof of the present invention,

[0523] (b) measuring with a photometer the absorbance of the sample to determine the total amount of hemoglobin in the sample,

[0524] (c) adding to the sample a polyhapten comprising at least one binding site for the antibody or antigen binding fragment thereof of the present invention, and

[0525] (d) measuring with a photometer the absorbance of the sample to determine the amount of HbAlc in the sample. All embodiments mentioned for any of the first to twenty-second and / or twenty -third aspect of the invention apply for the twenty-fourth aspect of the invention and vice versa.

[0526] The ratio of HbAlc, i.e. the amount of HbAlc in a sample compared to the amount of total hemoglobin in the same sample, is an indicator for the exposure of hemoglobin to constant high blood glucose levels and is therefore a valuable biomarker for the diagnosis of diabetes.

[0527] In embodiments, the absorbance in step (b) is measured at a wavelength of about 500 to about 650 nm, in particular 546 nm.

[0528] In embodiments, the absorbance in step (d) is measured at a wavelength of about 340 to about 600 nm, in particular 340 nm.

[0529] In embodiments, the absorbance measured in step (d) is inversely proportional to the concentration of HbAlc in the sample.

[0530] In embodiments, the sample is whole blood, which has been hemolysed before adding the antibody in step (a).

[0531] In embodiments, the antibody has been added in step (a) to a final concentration of about 0.025 to 10 mg / ml.

[0532] In embodiments, the polyhapten has been added in step (c) to a final concentration of about 2.5 to 50 μg / ml.

[0533] In embodiments, the method is a turbidimetric inhibition immunoassay.

[0534] In embodiments, the method has a coefficient of variation of 1.5 % or lower, when using a sample comprising a normal physiological concentration of HbAlc.

[0535] In embodiments, the method has a coefficient of variation of 0.8 % or lower, when using a sample comprising a normal physiological concentration of HbAlc.

[0536] In embodiments, the sample is comprising HbAlc at a concentration lower than 6.0 %, wherein the percentage is the amount of HbAlc compared to the total amount of hemoglobin. In embodiments, the method has a coefficient of variation of 1.5 % or lower, when using a sample comprising an abnormal physiological concentration of HbAlc. In specific embodiments, the method has a coefficient of variation of 1.2 % or lower, when using a sample comprising an abnormal physiological concentration of HbAlc.

[0537] In embodiments, the sample is comprising HbAlc at a concentration higher than or equal to 6.0 %, wherein the percentage is the amount of HbAlc compared to the total amount of hemoglobin.

[0538] In further embodiments, the present invention relates to the following items:

[0539] 1. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof.

[0540] 2. The antibody or antigen binding fragment thereof of item 1, wherein the variant of SEQ ID NO: 2 is having at least 85 % sequence identity to SEQ ID NO: 2.

[0541] 3. The antibody or antigen binding fragment thereof of item 1 or 2, wherein the variant of SEQ ID NO: 2 is having at least 90 % sequence identity to SEQ ID NO: 2.

[0542] 4. The antibody or antigen binding fragment thereof of any one of items 1 to 3, wherein the variant of SEQ ID NO: 2 is having at least 95 % sequence identity to SEQ ID NO: 2.

[0543] 5. The antibody or antigen binding fragment thereof of any one of items 1 to 4, wherein the variant of SEQ ID NO: 3 is having at least 85 % sequence identity to SEQ ID NO: 3.

[0544] 6. The antibody or antigen binding fragment thereof of any one of items 1 to 5, wherein the variant of SEQ ID NO: 3 is having at least 90 % sequence identity to SEQ ID NO: 3.

[0545] 7. The antibody or antigen binding fragment thereof of any one of items 1 to 6, wherein the variant of SEQ ID NO: 3 is having at least 95 % sequence identity to SEQ ID NO: 3.

[0546] 8. The antibody or antigen binding fragment thereof of any one of items 1 to 7, wherein the variant of SEQ ID NO: 4 is having at least 85 % sequence identity to SEQ ID NO: 4. 9. The antibody or antigen binding fragment thereof of any one of items 1 to 8, wherein the variant of SEQ ID NO: 4 is having at least 90 % sequence identity to SEQ ID NO: 4.

[0547] 10. The antibody or antigen binding fragment thereof of any one of items 1 to 9, wherein the variant of SEQ ID NO: 4 is having at least 95 % sequence identity to SEQ ID NO: 4.

[0548] 11. The antibody or antigen binding fragment thereof of any one of items 1 to 10, wherein the variant of SEQ ID NO: 6 is having at least 85 % sequence identity to SEQ ID NO: 6.

[0549] 12. The antibody or antigen binding fragment thereof of any one of items 1 to 11, wherein the variant of SEQ ID NO: 6 is having at least 90 % sequence identity to SEQ ID NO: 6.

[0550] 13. The antibody or antigen binding fragment thereof of any one of items 1 to 12, wherein the variant of SEQ ID NO: 6 is having at least 95 % sequence identity to SEQ ID NO: 6.

[0551] 14. The antibody or antigen binding fragment thereof of any one of items 1 to 13, wherein the variant of SEQ ID NO: 2 has

[0552] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19, and / or

[0553] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0554] 15. The antibody or antigen binding fragment thereof of item 14, wherein the variant of SEQ ID NO: 2 has

[0555] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0556] 16. The antibody or antigen binding fragment thereof of any one of items 1 to 14, wherein the variant of SEQ ID NO: 3 has

[0557] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0558] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0559] 17. The antibody or antigen binding fragment thereof of any one of items 1 to 16, wherein the variant of SEQ ID NO: 3 has

[0560] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0561] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0562] 18. The antibody or antigen binding fragment thereof of any one of items 1 to 17, wherein the variant of SEQ ID NO: 4 has

[0563] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0564] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0565] 19. The antibody or antigen binding fragment thereof of item 18, wherein the variant of SEQ ID NO: 4 has

[0566] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10. 20. The antibody or antigen binding fragment thereof of any one of items 1 to 18, wherein the variant of SEQ ID NO: 6 has

[0567] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0568] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0569] 21. The antibody or antigen binding fragment thereof of any one of items 1 to 20, wherein the variant of SEQ ID NO: 6 has

[0570] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0571] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0572] 22. The antibody or antigen binding fragment thereof of item 20 or 21, wherein the variant of SEQ ID NO: 6 has

[0573] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0574] 23. The antibody or antigen binding fragment thereof of any one of items 20 to 22, wherein the variant of SEQ ID NO: 6 has

[0575] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0576] 24. The antibody or antigen binding fragment thereof of any one of items 1 to 23, wherein the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof with one amino acid substitution. 25. The antibody or antigen binding fragment thereof of any one of items 1 to 24, wherein the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 2, a CDR-H3 as shown in SEQ ID NO: 3, a CDR-L1 as shown in SEQ ID NO: 4, and a CDR-L3 as shown in SEQ ID NO: 6.

[0577] 26. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof.

[0578] 27. The antibody or antigen binding fragment thereof of item 26, wherein the variant of SEQ ID NO: 8 is having at least 85 % sequence identity to SEQ ID NO: 8.

[0579] 28. The antibody or antigen binding fragment thereof of item 26 or 27, wherein the variant of SEQ ID NO: 8 is having at least 90 % sequence identity to SEQ ID NO: 8.

[0580] 29. The antibody or antigen binding fragment thereof of any one of items 26 to 28, wherein the variant of SEQ ID NO: 8 is having at least 95 % sequence identity to SEQ ID NO: 8.

[0581] 30. The antibody or antigen binding fragment thereof of any one of items 26 to 29, wherein the variant of SEQ ID NO: 9 is having at least 85 % sequence identity to SEQ ID NO: 9.

[0582] 31. The antibody or antigen binding fragment thereof of any one of items 26 to 30, wherein the variant of SEQ ID NO: 9 is having at least 90 % sequence identity to SEQ ID NO: 9.

[0583] 32. The antibody or antigen binding fragment thereof of any one of items 26 to 31, wherein the variant of SEQ ID NO: 9 is having at least 95 % sequence identity to SEQ ID NO: 9.

[0584] 33. The antibody or antigen binding fragment thereof of any one of items 26 to 32, wherein the variant of SEQ ID NO: 10 is having at least 85 % sequence identity to SEQ ID NO: 10.

[0585] 34. The antibody or antigen binding fragment thereof of any one of items 26 to 33, wherein the variant of SEQ ID NO: 10 is having at least 90 % sequence identity to SEQ ID NO: 10.

[0586] 35. The antibody or antigen binding fragment thereof of any one of items 26 to 34, wherein the variant of SEQ ID NO: 10 is having at least 95 % sequence identity to SEQ ID NO: 10. 36. The antibody or antigen binding fragment thereof of any one of items 26 to 35, wherein the variant of SEQ ID NO: 12 is having at least 85 % sequence identity to SEQ ID NO: 12.

[0587] 37. The antibody or antigen binding fragment thereof of any one of items 26 to 36, wherein the variant of SEQ ID NO: 12 is having at least 90 % sequence identity to SEQ ID NO: 12.

[0588] 38. The antibody or antigen binding fragment thereof of any one of items 26 to 37, wherein the variant of SEQ ID NO: 12 is having at least 95 % sequence identity to SEQ ID NO: 12.

[0589] 39. The antibody or antigen binding fragment thereof of any one of items 26 to 38, wherein the variant of SEQ ID NO: 8 has

[0590] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19;

[0591] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T; and / or

[0592] (iii) deletion of at most three amino acids.

[0593] 40. The antibody or antigen binding fragment thereof of item 39, wherein the variant of SEQ ID NO: 8 has

[0594] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0595] 41. The antibody or antigen binding fragment thereof of any one of items 26 to 40, wherein the variant of SEQ ID NO: 9 has

[0596] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0597] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0598] 42. The antibody or antigen binding fragment thereof of any one of items 26 to 41, wherein the variant of SEQ ID NO: 9 has

[0599] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0600] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0601] 43. The antibody or antigen binding fragment thereof of any one of items 26 to 42, wherein the variant of SEQ ID NO: 10 has

[0602] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0603] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0604] 44. The antibody or antigen binding fragment thereof of any one of items 26 to 43, wherein the variant of SEQ ID NO: 10 has

[0605] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0606] 45. The antibody or antigen binding fragment thereof of any one of items 26 to 44, wherein the variant of SEQ ID NO: 12 has (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0607] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0608] 46. The antibody or antigen binding fragment thereof of any one of items 26 to 45, wherein the variant of SEQ ID NO: 12 has

[0609] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0610] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0611] 47. The antibody or antigen binding fragment thereof of item 45 or 46, wherein the variant of SEQ ID NO: 12 has

[0612] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0613] 48. The antibody or antigen binding fragment thereof of any one of items 45 to 47, wherein the variant of SEQ ID NO: 12 has

[0614] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0615] 49. The antibody or antigen binding fragment thereof of any one of items 26 to 48, wherein the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof with one amino acid substitution.

[0616] 50. The antibody or antigen binding fragment thereof of any one of items 26 to 49, wherein the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 8, a CDR-H3 as shown in SEQ ID NO: 9, a CDR-L1 as shown in SEQ ID NO: 10, and a CDR-L3 as shown in SEQ ID NO: 12.

[0617] 51. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof.

[0618] 52. The antibody or antigen binding fragment thereof of item 51, wherein the variant of SEQ ID NO: 14 is having at least 85 % sequence identity to SEQ ID NO: 14.

[0619] 53. The antibody or antigen binding fragment thereof of item 51 or 52, wherein the variant of SEQ ID NO: 14 is having at least 90 % sequence identity to SEQ ID NO: 14.

[0620] 54. The antibody or antigen binding fragment thereof of any one of items 51 to 53, wherein the variant of SEQ ID NO: 14 is having at least 95 % sequence identity to SEQ ID NO: 14.

[0621] 55. The antibody or antigen binding fragment thereof of any one of items 51 to 54, wherein the variant of SEQ ID NO: 15 is having at least 85 % sequence identity to SEQ ID NO: 15.

[0622] 56. The antibody or antigen binding fragment thereof of any one of items 51 to 55, wherein the variant of SEQ ID NO: 15 is having at least 90 % sequence identity to SEQ ID NO: 15.

[0623] 57. The antibody or antigen binding fragment thereof of any one of items 51 to 56, wherein the variant of SEQ ID NO: 15 is having at least 95 % sequence identity to SEQ ID NO: 15.

[0624] 58. The antibody or antigen binding fragment thereof of any one of items 51 to 57, wherein the variant of SEQ ID NO: 16 is having at least 85 % sequence identity to SEQ ID NO: 16.

[0625] 59. The antibody or antigen binding fragment thereof of any one of items 51 to 58, wherein the variant of SEQ ID NO: 16 is having at least 90 % sequence identity to SEQ ID NO: 16.

[0626] 60. The antibody or antigen binding fragment thereof of any one of items 51 to 59, wherein the variant of SEQ ID NO: 16 is having at least 95 % sequence identity to SEQ ID NO: 16.

[0627] 61. The antibody or antigen binding fragment thereof of any one of items 51 to 60, wherein the variant of SEQ ID NO: 18 is having at least 85 % sequence identity to SEQ ID NO: 18. 62. The antibody or antigen binding fragment thereof of any one of items 51 to 61, wherein the variant of SEQ ID NO: 18 is having at least 90 % sequence identity to SEQ ID NO: 18.

[0628] 63. The antibody or antigen binding fragment thereof of any one of items 51 to 62, wherein the variant of SEQ ID NO: 18 is having at least 95 % sequence identity to SEQ ID NO: 18.

[0629] 64. The antibody or antigen binding fragment thereof of any one of items 51 to 63, wherein the variant of SEQ ID NO: 14 has

[0630] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 16;

[0631] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0632] 65. The antibody or antigen binding fragment thereof of any one of items 51 to 64, wherein the variant of SEQ ID NO: 14 has

[0633] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 16.

[0634] 66. The antibody or antigen binding fragment thereof of any one of items 51 to 65, wherein the variant of SEQ ID NO: 15 has

[0635] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0636] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0637] 67. The antibody or antigen binding fragment thereof of any one of items 51 to 66, wherein the variant of SEQ ID NO: 15 has (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0638] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0639] 68. The antibody or antigen binding fragment thereof of any one of items 51 to 67, wherein the variant of SEQ ID NO: 16 has

[0640] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0641] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0642] 69. The antibody or antigen binding fragment thereof of item 68, wherein the variant of SEQ ID NO: 16 has

[0643] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0644] 70. The antibody or antigen binding fragment thereof of any one of items 51 to 69, wherein the variant of SEQ ID NO: 18 has

[0645] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0646] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W. 71. The antibody or antigen binding fragment thereof of any one of items 51 to 70, wherein the variant of SEQ ID NO: 18 has

[0647] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0648] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0649] 72. The antibody or antigen binding fragment thereof of item 70 or 71, wherein the variant of SEQ ID NO: 18 has

[0650] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0651] 73. The antibody or antigen binding fragment thereof of any one of items 70 to 72, wherein the variant of SEQ ID NO: 18 has

[0652] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0653] 74. The antibody or antigen binding fragment thereof of any one of items 51 to 73, wherein the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof with one amino acid substitution.

[0654] 75. The antibody or antigen binding fragment thereof of any one of items 51 to 74, wherein the antibody or antigen binding fragment thereof comprises a CDR-H2 as shown in SEQ ID NO: 14, a CDR-H3 as shown in SEQ ID NO: 15, a CDR-L1 as shown in SEQ ID NO: 16 and a CDR-L3 as shown in SEQ ID NO: 18.

[0655] 76. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 1 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 5 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof.

[0656] 77. The antibody or antigen binding fragment thereof of item 76, wherein the variant of SEQ ID NO: 1 is having at least 85 % sequence identity to SEQ ID NO: 1.

[0657] 78. The antibody or antigen binding fragment thereof of item 76 or 77, wherein the variant of SEQ ID NO: 1 is having at least 90 % sequence identity to SEQ ID NO: 1.

[0658] 79. The antibody or antigen binding fragment thereof of any one of items 76 to 78, wherein the variant of SEQ ID NO: 1 is having at least 95 % sequence identity to SEQ ID NO: 1.

[0659] 80. The antibody or antigen binding fragment thereof of any one of items 76 to 79, wherein the variant of SEQ ID NO: 2 is having at least 85 % sequence identity to SEQ ID NO: 2.

[0660] 81. The antibody or antigen binding fragment thereof of any one of items 76 to 80, wherein the variant of SEQ ID NO: 2 is having at least 90 % sequence identity to SEQ ID NO: 2.

[0661] 82. The antibody or antigen binding fragment thereof of any one of items 76 to 81, wherein the variant of SEQ ID NO: 2 is having at least 95 % sequence identity to SEQ ID NO: 2.

[0662] 83. The antibody or antigen binding fragment thereof of any one of items 76 to 82, wherein the variant of SEQ ID NO: 3 is having at least 85 % sequence identity to SEQ ID NO: 3.

[0663] 84. The antibody or antigen binding fragment thereof of any one of items 76 to 83, wherein the variant of SEQ ID NO: 3 is having at least 90 % sequence identity to SEQ ID NO: 3.

[0664] 85. The antibody or antigen binding fragment thereof of any one of items 76 to 84, wherein the variant of SEQ ID NO: 3 is having at least 95 % sequence identity to SEQ ID NO: 3.

[0665] 86. The antibody or antigen binding fragment thereof of any one of items 76 to 85, wherein the variant of SEQ ID NO: 4 is having at least 85 % sequence identity to SEQ ID NO: 4.

[0666] 87. The antibody or antigen binding fragment thereof of any one of items 76 to 86, wherein the variant of SEQ ID NO: 4 is having at least 90 % sequence identity to SEQ ID NO: 4. 88. The antibody or antigen binding fragment thereof of any one of items 76 to 87, wherein the variant of SEQ ID NO: 4 is having at least 95 % sequence identity to SEQ ID NO: 4.

[0667] 89. The antibody or antigen binding fragment thereof of any one of items 76 to 88, wherein the variant of SEQ ID NO: 5 is having at least 85 % sequence identity to SEQ ID NO: 5.

[0668] 90. The antibody or antigen binding fragment thereof of any one of items 76 to 89, wherein the variant of SEQ ID NO: 5 is having at least 90 % sequence identity to SEQ ID NO: 5.

[0669] 91. The antibody or antigen binding fragment thereof of any one of items 76 to 90, wherein the variant of SEQ ID NO: 5 is having at least 95 % sequence identity to SEQ ID NO: 5.

[0670] 92. The antibody or antigen binding fragment thereof of any one of items 76 to 91, wherein the variant of SEQ ID NO: 6 is having at least 85 % sequence identity to SEQ ID NO: 6.

[0671] 93. The antibody or antigen binding fragment thereof of any one of items 76 to 92, wherein the variant of SEQ ID NO: 6 is having at least 90 % sequence identity to SEQ ID NO: 6.

[0672] 94. The antibody or antigen binding fragment thereof of any one of items 76 to 93, wherein the variant of SEQ ID NO: 6 is having at least 95 % sequence identity to SEQ ID NO: 6.

[0673] 95. The antibody or antigen binding fragment thereof of any one of items 76 to 94, wherein the variant of SEQ ID NO: 1 has

[0674] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0675] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0676] 96. The antibody or antigen binding fragment thereof of any one of items 76 to 95, wherein the variant of SEQ ID NO: 1 has

[0677] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0678] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W. 97. The antibody or antigen binding fragment thereof of any one of items 76 to 96, wherein the variant of SEQ ID NO: 2 has

[0679] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19, and / or

[0680] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0681] 98. The antibody or antigen binding fragment thereof of any one of items 76 to 97, wherein the variant of SEQ ID NO: 2 has

[0682] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0683] 99. The antibody or antigen binding fragment thereof of any one of items 76 to 98, wherein the variant of SEQ ID NO: 3 has

[0684] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0685] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0686] 100. The antibody or antigen binding fragment thereof of any one of items 76 to 99, wherein the variant of SEQ ID NO: 3 has

[0687] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0688] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0689] 101. The antibody or antigen binding fragment thereof of any one of items 76 to 100, wherein the variant of SEQ ID NO: 4 has

[0690] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0691] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0692] 102. The antibody or antigen binding fragment thereof of any one of items 76 to 101, wherein the variant of SEQ ID NO: 4 has

[0693] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0694] 103. The antibody or antigen binding fragment thereof of any one of items 76 to 102, wherein the variant of SEQ ID NO: 6 has

[0695] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0696] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0697] 104. The antibody or antigen binding fragment thereof of any one of items 76 to 103, wherein the variant of SEQ ID NO: 6 has

[0698] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0699] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0700] 105. The antibody or antigen binding fragment thereof of any one of items 76 to 104, wherein the variant of SEQ ID NO: 6 has

[0701] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0702] 106. The antibody or antigen binding fragment thereof of any one of items 76 to 105, wherein the variant of SEQ ID NO: 6 has

[0703] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0704] 107. The antibody or antigen binding fragment thereof of any one of items 76 to 106, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 1 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 5 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof with one amino acid substitution.

[0705] 108. The antibody or antigen binding fragment thereof of any one of items 76 to 107, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 1, a CDR-H2 as shown in SEQ ID NO: 2, a CDR-H3 as shown in SEQ ID NO: 3, a CDR-L1 as shown in SEQ ID NO: 4, a CDR-L2 as shown in SEQ ID NO: 5 and a CDR-L3 as shown in SEQ ID NO: 6.

[0706] 109. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 7 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 11 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof. 110. The antibody or antigen binding fragment thereof of item 109, wherein the variant of SEQ ID NO: 7 is having at least 85 % sequence identity to SEQ ID NO: 7.

[0707] 111. The antibody or antigen binding fragment thereof of item 109 or 110, wherein the variant of SEQ ID NO: 7 is having at least 90 % sequence identity to SEQ ID NO: 7.

[0708] 112. The antibody or antigen binding fragment thereof of any one of items 109 to 111, wherein the variant of SEQ ID NO: 7 is having at least 95 % sequence identity to SEQ ID NO: 7.

[0709] 113. The antibody or antigen binding fragment thereof of any one of items 109 to 112, wherein the variant of SEQ ID NO: 8 is having at least 85 % sequence identity to SEQ ID NO: 8.

[0710] 114. The antibody or antigen binding fragment thereof of any one of items 109 to 113, wherein the variant of SEQ ID NO: 8 is having at least 90 % sequence identity to SEQ ID NO: 8.

[0711] 115. The antibody or antigen binding fragment thereof of any one of items 109 to 114, wherein the variant of SEQ ID NO: 8 is having at least 95 % sequence identity to SEQ ID NO: 8.

[0712] 116. The antibody or antigen binding fragment thereof of any one of items 109 to 115, wherein the variant of SEQ ID NO: 9 is having at least 85 % sequence identity to SEQ ID NO: 9.

[0713] 117. The antibody or antigen binding fragment thereof of any one of items 109 to 116, wherein the variant of SEQ ID NO: 9 is having at least 90 % sequence identity to SEQ ID NO: 9.

[0714] 118. The antibody or antigen binding fragment thereof of any one of items 109 to 117, wherein the variant of SEQ ID NO: 9 is having at least 95 % sequence identity to SEQ ID NO: 9.

[0715] 119. The antibody or antigen binding fragment thereof of any one of items 109 to 118, wherein the variant of SEQ ID NO: 10 is having at least 85 % sequence identity to SEQ ID NO: 10.

[0716] 120. The antibody or antigen binding fragment thereof of any one of items 109 to 119, wherein the variant of SEQ ID NO: 10 is having at least 90 % sequence identity to SEQ ID NO: 10.

[0717] 121. The antibody or antigen binding fragment thereof of any one of items 109 to 120, wherein the variant of SEQ ID NO: 10 is having at least 95 % sequence identity to SEQ ID NO: 10. 122. The antibody or antigen binding fragment thereof of any one of items 109 to 121, wherein the variant of SEQ ID NO: 11 is having at least 85 % sequence identity to SEQ ID NO: 11.

[0718] 123. The antibody or antigen binding fragment thereof of any one of items 109 to 122, wherein the variant of SEQ ID NO: 11 is having at least 90 % sequence identity to SEQ ID NO: 11.

[0719] 124. The antibody or antigen binding fragment thereof of any one of items 109 to 123, wherein the variant of SEQ ID NO: 11 is having at least 95 % sequence identity to SEQ ID NO: 11.

[0720] 125. The antibody or antigen binding fragment thereof of any one of items 109 to 124, wherein the variant of SEQ ID NO: 12 is having at least 85 % sequence identity to SEQ ID NO: 12.

[0721] 126. The antibody or antigen binding fragment thereof of any one of items 109 to 125, wherein the variant of SEQ ID NO: 12 is having at least 90 % sequence identity to SEQ ID NO: 12.

[0722] 127. The antibody or antigen binding fragment thereof of any one of items 109 to 126, wherein the variant of SEQ ID NO: 12 is having at least 95 % sequence identity to SEQ ID NO: 12.

[0723] 128. The antibody or antigen binding fragment thereof of any one of items 109 to 127, wherein the variant of SEQ ID NO: 7 has

[0724] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0725] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0726] 129. The antibody or antigen binding fragment thereof of any one of items 109 to 128, wherein the variant of SEQ ID NO: 7 has

[0727] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0728] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0729] 130. The antibody or antigen binding fragment thereof of any one of items 109 to 129, wherein the variant of SEQ ID NO: 8 has

[0730] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19; (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T; and / or

[0731] (iii) deletion of at most three amino acids.

[0732] 131. The antibody or antigen binding fragment thereof of item 130, wherein the variant of SEQ ID NO: 8 has

[0733] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0734] 132. The antibody or antigen binding fragment thereof of any one of items 109 to 131, wherein the variant of SEQ ID NO: 9 has

[0735] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0736] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0737] 133. The antibody or antigen binding fragment thereof of any one of items 109 to 132, wherein the variant of SEQ ID NO: 9 has

[0738] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0739] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0740] 134. The antibody or antigen binding fragment thereof of any one of items 109 to 133, wherein the variant of SEQ ID NO: 10 has

[0741] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0742] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0743] 135. The antibody or antigen binding fragment thereof of item 134, wherein the variant of SEQ ID NO: 10 has

[0744] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0745] 136. The antibody or antigen binding fragment thereof of any one of items 109 to 135, wherein the variant of SEQ ID NO: 12 has

[0746] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0747] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0748] 137. The antibody or antigen binding fragment thereof of any one of items 109 to 136, wherein the variant of SEQ ID NO: 12 has

[0749] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0750] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W. 138. The antibody or antigen binding fragment thereof of item 136 or 137, wherein the variant of SEQ ID NO: 12 has

[0751] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0752] 139. The antibody or antigen binding fragment thereof of any one of items 136 to 138, wherein the variant of SEQ ID NO: 12 has

[0753] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0754] 140. The antibody or antigen binding fragment thereof of any one of items 109 to 139, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 7 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 8 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 9 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 10 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 11 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 12 or variant thereof with one amino acid substitution.

[0755] 141. The antibody or antigen binding fragment thereof of any one of items 109 to 140, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 7, a CDR-H2 as shown in SEQ ID NO: 8, a CDR-H3 as shown in SEQ ID NO: 9, a CDR-L1 as shown in SEQ ID NO: 10, a CDR-L2 as shown in SEQ ID NO: 11 and a CDR-L3 as shown in SEQ ID NO: 12.

[0756] 142. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 13 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 17 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof.

[0757] 143. The antibody or antigen binding fragment thereof of item 142, wherein the variant of SEQ ID NO: 13 is having at least 85 % sequence identity to SEQ ID NO: 13.

[0758] 144. The antibody or antigen binding fragment thereof of item 142 or 143, wherein the variant of SEQ ID NO: 13 is having at least 90 % sequence identity to SEQ ID NO: 13. 145. The antibody or antigen binding fragment thereof of any one of items 142 to 144, wherein the variant of SEQ ID NO: 13 is having at least 95 % sequence identity to SEQ ID NO: 13.

[0759] 146. The antibody or antigen binding fragment thereof of any one of items 142 to 145, wherein the variant of SEQ ID NO: 14 is having at least 85 % sequence identity to SEQ ID NO: 14.

[0760] 147. The antibody or antigen binding fragment thereof of any one of items 142 to 146, wherein the variant of SEQ ID NO: 14 is having at least 90 % sequence identity to SEQ ID NO: 14.

[0761] 148. The antibody or antigen binding fragment thereof of any one of items 142 to 147, wherein the variant of SEQ ID NO: 14 is having at least 95 % sequence identity to SEQ ID NO: 14.

[0762] 149. The antibody or antigen binding fragment thereof of any one of items 142 to 148, wherein the variant of SEQ ID NO: 15 is having at least 85 % sequence identity to SEQ ID NO: 15.

[0763] 150. The antibody or antigen binding fragment thereof of any one of items 142 to 149, wherein the variant of SEQ ID NO: 15 is having at least 90 % sequence identity to SEQ ID NO: 15.

[0764] I51. The antibody or antigen binding fragment thereof of any one of items 142 to 150, wherein the variant of SEQ ID NO: 15 is having at least 95 % sequence identity to SEQ ID NO: 15.

[0765] 152. The antibody or antigen binding fragment thereof of any one of items 142 to I51, wherein the variant of SEQ ID NO: 16 is having at least 85 % sequence identity to SEQ ID NO: 16.

[0766] 153. The antibody or antigen binding fragment thereof of any one of items 142 to 152, wherein the variant of SEQ ID NO: 16 is having at least 90 % sequence identity to SEQ ID NO: 16.

[0767] 154. The antibody or antigen binding fragment thereof of any one of items 142 to 153, wherein the variant of SEQ ID NO: 16 is having at least 95 % sequence identity to SEQ ID NO: 16.

[0768] 155. The antibody or antigen binding fragment thereof of any one of items 142 to 154, wherein the variant of SEQ ID NO: 17 is having at least 85 % sequence identity to SEQ ID NO: 17.

[0769] 156. The antibody or antigen binding fragment thereof of any one of items 142 to 155, wherein the variant of SEQ ID NO: 17 is having at least 90 % sequence identity to SEQ ID NO: 17. 157. The antibody or antigen binding fragment thereof of any one of items 142 to 156, wherein the variant of SEQ ID NO: 17 is having at least 95 % sequence identity to SEQ ID NO: 17.

[0770] 158. The antibody or antigen binding fragment thereof of any one of items 142 to 157, wherein the variant of SEQ ID NO: 18 is having at least 85 % sequence identity to SEQ ID NO: 18.

[0771] 159. The antibody or antigen binding fragment thereof of any one of items 142 to 158, wherein the variant of SEQ ID NO: 18 is having at least 90 % sequence identity to SEQ ID NO: 18.

[0772] 160. The antibody or antigen binding fragment thereof of any one of items 142 to 159, wherein the variant of SEQ ID NO: 18 is having at least 95 % sequence identity to SEQ ID NO: 18.

[0773] 161. The antibody or antigen binding fragment thereof of any one of items 142 to 160, wherein the variant of SEQ ID NO: 13 has

[0774] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0775] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0776] 162. The antibody or antigen binding fragment thereof of any one of items 142 to 161, wherein the variant of SEQ ID NO: 13 has

[0777] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0778] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0779] 163. The antibody or antigen binding fragment thereof of any one of items 142 to 162, wherein the variant of SEQ ID NO: 14 has

[0780] (i) at most four, in particular two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 16;

[0781] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T. 164. The antibody or antigen binding fragment thereof of any one of items 142 to 163, wherein the variant of SEQ ID NO: 14 has

[0782] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 16.

[0783] 165. The antibody or antigen binding fragment thereof of any one of items 142 to 163, wherein the variant of SEQ ID NO: 15 has

[0784] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0785] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0786] 166. The antibody or antigen binding fragment thereof of any one of items 142 to 165, wherein the variant of SEQ ID NO: 15 has

[0787] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0788] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W. 167. The antibody or antigen binding fragment thereof of any one of items 142 to 166, wherein the variant of SEQ ID NO: 16 has

[0789] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0790] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0791] 168. The antibody or antigen binding fragment thereof of item 167, wherein the variant of SEQ ID NO: 16 has

[0792] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0793] 169. The antibody or antigen binding fragment thereof of any one of items 142 to 168, wherein the variant of SEQ ID NO: 18 has

[0794] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0795] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0796] 170. The antibody or antigen binding fragment thereof of any one of items 142 to 169, wherein the variant of SEQ ID NO: 18 has

[0797] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0798] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0799] 171. The antibody or antigen binding fragment thereof of item 169 or 170, wherein the variant of SEQ ID NO: 18 has

[0800] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12. 172. The antibody or antigen binding fragment thereof of any one of items 169 to 171, wherein the variant of SEQ ID NO: 18 has

[0801] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0802] 173. The antibody or antigen binding fragment thereof of any one of items 142 to 172, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 13 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 14 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 15 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 16 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 17 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 18 or variant thereof with one amino acid substitution.

[0803] 174. The antibody or antigen binding fragment thereof of any one of items 142 to 173, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 13, a CDR-H2 as shown in SEQ ID NO: 14, a CDR-H3 as shown in SEQ ID NO: 15, a CDR-L1 as shown in SEQ ID NO: 16, a CDR-L2 as shown in SEQ ID NO: 17 and a CDR-L3 as shown in SEQ ID NO: 18.

[0804] 175. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H1 as shown in SEQ ID NO: 19 or variant thereof, a CDR-H2 as shown in SEQ ID NO: 20 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 21 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 22 or variant thereof, a CDR-L2 as shown in SEQ ID NO: 23 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 24 or variant thereof.

[0805] 176. The antibody or antigen binding fragment thereof of item 175, wherein the variant of SEQ ID NO: 19 is having at least 85 % sequence identity to SEQ ID NO: 19.

[0806] 177. The antibody or antigen binding fragment thereof of item 175 or 176, wherein the variant of SEQ ID NO: 19 is having at least 90 % sequence identity to SEQ ID NO: 19.

[0807] 178. The antibody or antigen binding fragment thereof of any one of items 175 to 177, wherein the variant of SEQ ID NO: 19 is having at least 95 % sequence identity to SEQ ID NO: 19. 179. The antibody or antigen binding fragment thereof of any one of items 175 to 178, wherein the variant of SEQ ID NO: 20 is having at least 85 % sequence identity to SEQ ID NO: 20.

[0808] 180. The antibody or antigen binding fragment thereof of any one of items 175 to 179, wherein the variant of SEQ ID NO: 20 is having at least 90 % sequence identity to SEQ ID NO: 20.

[0809] 181. The antibody or antigen binding fragment thereof of any one of items 175 to 180, wherein the variant of SEQ ID NO: 20 is having at least 95 % sequence identity to SEQ ID NO: 20.

[0810] 182. The antibody or antigen binding fragment thereof of any one of items 175 to 181, wherein the variant of SEQ ID NO: 21 is having at least 85 % sequence identity to SEQ ID NO: 21.

[0811] 183. The antibody or antigen binding fragment thereof of any one of items 175 to 182, wherein the variant of SEQ ID NO: 21 is having at least 90 % sequence identity to SEQ ID NO: 21.

[0812] 184. The antibody or antigen binding fragment thereof of any one of items 175 to 183, wherein the variant of SEQ ID NO: 21 is having at least 95 % sequence identity to SEQ ID NO: 21.

[0813] 185. The antibody or antigen binding fragment thereof of any one of items 175 to 184, wherein the variant of SEQ ID NO: 22 is having at least 85 % sequence identity to SEQ ID NO: 22.

[0814] 186. The antibody or antigen binding fragment thereof of any one of items 175 to 185, wherein the variant of SEQ ID NO: 22 is having at least 90 % sequence identity to SEQ ID NO: 22.

[0815] 187. The antibody or antigen binding fragment thereof of any one of items 175 to 186, wherein the variant of SEQ ID NO: 22 is having at least 95 % sequence identity to SEQ ID NO: 22.

[0816] 188. The antibody or antigen binding fragment thereof of any one of items 175 to 187, wherein the variant of SEQ ID NO: 23 is having at least 85 % sequence identity to SEQ ID NO: 23.

[0817] 189. The antibody or antigen binding fragment thereof of any one of items 175 to 188, wherein the variant of SEQ ID NO: 23 is having at least 90 % sequence identity to SEQ ID NO: 23.

[0818] 190. The antibody or antigen binding fragment thereof of any one of items 175 to 189, wherein the variant of SEQ ID NO: 23 is having at least 95 % sequence identity to SEQ ID NO: 23. 191. The antibody or antigen binding fragment thereof of any one of items 175 to 190, wherein the variant of SEQ ID NO: 24 is having at least 85 % sequence identity to SEQ ID NO: 24.

[0819] 192. The antibody or antigen binding fragment thereof of any one of items 175 to 191, wherein the variant of SEQ ID NO: 24 is having at least 90 % sequence identity to SEQ ID NO: 24.

[0820] 193. The antibody or antigen binding fragment thereof of any one of items 175 to 192, wherein the variant of SEQ ID NO: 24 is having at least 95 % sequence identity to SEQ ID NO: 24.

[0821] 194. The antibody or antigen binding fragment thereof of any one of items 175 to 193, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 19 or variant thereof with one amino acid substitution, a CDR-H2 as shown in SEQ ID NO: 20 or variant thereof with one amino acid substitution, a CDR-H3 as shown in SEQ ID NO: 21 or variant thereof with one amino acid substitution, a CDR-L1 as shown in SEQ ID NO: 22 or variant thereof with one amino acid substitution, a CDR-L2 as shown in SEQ ID NO: 23 or variant thereof with one amino acid substitution, and a CDR-L3 as shown in SEQ ID NO: 24 or variant thereof with one amino acid substitution.

[0822] 195. The antibody or antigen binding fragment thereof of any one of items 175 to 194, wherein the antibody or antigen binding fragment thereof comprises a CDR-H1 as shown in SEQ ID NO: 19, a CDR-H2 as shown in SEQ ID NO: 20, a CDR-H3 as shown in SEQ ID NO: 21, a CDR-L1 as shown in SEQ ID NO: 22, a CDR-L2 as shown in SEQ ID NO: 23 and a CDR-L3 as shown in SEQ ID NO: 24.

[0823] 196. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: 25 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 26 or variant thereof.

[0824] 197. The antibody or antigen binding fragment thereof of item 196, wherein the variant of SEQ ID NO: 25 is having at least 85 % sequence identity.

[0825] 198. The antibody or antigen binding fragment thereof of item 196 or 197, wherein the variant of SEQ ID NO: 25 is having at least 90 % sequence identity.

[0826] 199. The antibody or antigen binding fragment thereof of any one of items 196 to 198, wherein the variant of SEQ ID NO: 25 is having at least 90 % sequence identity. 200. The antibody or antigen binding fragment thereof of any one of items 196 to 199, wherein the variant of SEQ ID NO: 26 is having at least 85 % sequence identity.

[0827] 201. The antibody or antigen binding fragment thereof of any one of items 196 to 200, wherein the variant of SEQ ID NO: 26 is having at least 90 % sequence identity.

[0828] 202. The antibody or antigen binding fragment thereof of any one of items 196 to 201, wherein the variant of SEQ ID NO: 26 is having at least 95 % sequence identity.

[0829] 203. The antibody or antigen binding fragment thereof of any one of items 196 to 202, wherein the heavy chain variable domain comprises

[0830] a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 1 or a variant thereof,

[0831] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 2 or a variant thereof, and

[0832] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 3 or a variant thereof; and / or

[0833] the light chain variable domain comprises

[0834] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 4 or a variant thereof,

[0835] a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 5 or a variant thereof, and

[0836] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 6 or a variant thereof.

[0837] 204. The antibody or antigen binding fragment thereof of item 203, wherein the variant of SEQ ID NO: 1 has

[0838] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0839] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0840] 205. The antibody or antigen binding fragment thereof of item 203 or 204, wherein the variant of SEQ ID NO: 1 has (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0841] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0842] 206. The antibody or antigen binding fragment thereof of any one of items 203 to 205, wherein the variant of SEQ ID NO: 2 has

[0843] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19, and / or

[0844] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0845] 207. The antibody or antigen binding fragment thereof of any one of items 203 to 206, wherein the variant of SEQ ID NO: 2 has

[0846] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0847] 208. The antibody or antigen binding fragment thereof of any one of items 203 to 207, wherein the variant of SEQ ID NO: 3 has

[0848] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0849] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0850] 209. The antibody or antigen binding fragment thereof of any one of items 203 to 208, wherein the variant of SEQ ID NO: 3 has (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0851] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0852] 210. The antibody or antigen binding fragment thereof of any one of items 203 to 209, wherein the variant of SEQ ID NO: 4 has

[0853] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0854] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0855] 211. The antibody or antigen binding fragment thereof of item 210, wherein the variant of SEQ ID NO: 4 has

[0856] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0857] 212. The antibody or antigen binding fragment thereof of any one of items 203 to 211, wherein the variant of SEQ ID NO: 6 has

[0858] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0859] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W. 213. The antibody or antigen binding fragment thereof of any one of items 203 to 212, wherein the variant of SEQ ID NO: 6 has

[0860] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0861] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0862] 214. The antibody or antigen binding fragment thereof of item 212 or 213, wherein the variant of SEQ ID NO: 6 has

[0863] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0864] 215. The antibody or antigen binding fragment thereof of any one of items 212 to 214, wherein the variant of SEQ ID NO: 6 has

[0865] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0866] 216. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: SEQ ID NO: 27 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 28 or variant thereof.

[0867] 217. The antibody or antigen binding fragment thereof of item 216, wherein the variant of SEQ ID NO: 27 is having at least 85 % sequence identity.

[0868] 218. The antibody or antigen binding fragment thereof of item 216 or 217, wherein the variant of SEQ ID NO: 27 is having at least 90 % sequence identity.

[0869] 219. The antibody or antigen binding fragment thereof of any one of items 216 to 218, wherein the variant of SEQ ID NO: 27 is having at least 95 % sequence identity.

[0870] 220. The antibody or antigen binding fragment thereof of any one of items 216 to 219, wherein the variant of SEQ ID NO: 28 is having at least 85 % sequence identity. 221. The antibody or antigen binding fragment thereof of any one of items 216 to 220, wherein the variant of SEQ ID NO: 28 is having at least 90 % sequence identity.

[0871] 222. The antibody or antigen binding fragment thereof of any one of items 216 to 221, wherein the variant of SEQ ID NO: 28 is having at least 95 % sequence identity.

[0872] 223. The antibody or antigen binding fragment thereof of any one of items 216 to 222, wherein the heavy chain variable domain comprises

[0873] a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 7 or a variant thereof,

[0874] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 8 or a variant thereof, and

[0875] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 9 or a variant thereof; and / or

[0876] the light chain variable domain comprises

[0877] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 10 or a variant thereof,

[0878] a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 11 or a variant thereof, and

[0879] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 12 or a variant thereof.

[0880] 224. The antibody or antigen binding fragment thereof of item 223, wherein the variant of SEQ ID NO: 7 has

[0881] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0882] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0883] 225. The antibody or antigen binding fragment thereof of item 223 or 224, wherein the variant of SEQ ID NO: 7 has

[0884] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W.

[0885] 226. The antibody or antigen binding fragment thereof of any one of items 223 to 225, wherein the variant of SEQ ID NO: 8 has

[0886] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19;

[0887] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T; and / or

[0888] (iii) deletion of at most three amino acids.

[0889] 227. The antibody or antigen binding fragment thereof of item 226, wherein the variant of SEQ ID NO: 8 has

[0890] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 19.

[0891] 228. The antibody or antigen binding fragment thereof of any one of items 223 to 227, wherein the variant of SEQ ID NO: 9 has

[0892] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0893] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0894] 229. The antibody or antigen binding fragment thereof of any one of items 223 to 228, wherein the variant of SEQ ID NO: 9 has

[0895] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0896] 230. The antibody or antigen binding fragment thereof of any one of items 223 to 229, wherein the variant of SEQ ID NO: 10 has

[0897] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0898] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0899] 231. The antibody or antigen binding fragment thereof of item 230, wherein the variant of SEQ ID NO: 10 has

[0900] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0901] 232. The antibody or antigen binding fragment thereof of any one of items 223 to 231, wherein the variant of SEQ ID NO: 12 has

[0902] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0903] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0904] 233. The antibody or antigen binding fragment thereof of any one of items 223 to 232, wherein the variant of SEQ ID NO: 12 has (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0905] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0906] 234. The antibody or antigen binding fragment thereof of item 232 or 233, wherein the variant of SEQ ID NO: 12 has

[0907] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0908] 235. The antibody or antigen binding fragment thereof of any one of items 232 to 234, wherein the variant of SEQ ID NO: 12 has

[0909] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0910] 236. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: 29 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 30 or variant thereof.

[0911] 237. The antibody or antigen binding fragment thereof of item 236, wherein the variant of SEQ ID NO: 29 is having at least 85 % sequence identity.

[0912] 238. The antibody or antigen binding fragment thereof of item 236 or 237, wherein the variant of SEQ ID NO: 29 is having at least 90 % sequence identity.

[0913] 239. The antibody or antigen binding fragment thereof of any one of items 236 to 238, wherein the variant of SEQ ID NO: 29 is having at least 95 % sequence identity.

[0914] 240. The antibody or antigen binding fragment thereof of any one of items 236 to 239, wherein the variant of SEQ ID NO: 30 is having at least 85 % sequence identity.

[0915] 241. The antibody or antigen binding fragment thereof of any one of items 236 to 240, wherein the variant of SEQ ID NO: 30 is having at least 90 % sequence identity. 242. The antibody or antigen binding fragment thereof of any one of items 236 to 241, wherein the variant of SEQ ID NO: 30 is having at least 95 % sequence identity.

[0916] 243. The antibody or antigen binding fragment thereof of any one of items 236 to 242, wherein the heavy chain variable domain comprises

[0917] a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 13 or a variant thereof,

[0918] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 14 or a variant thereof, and

[0919] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 15 or a variant thereof; and / or

[0920] the light chain variable domain comprises

[0921] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 16 or a variant thereof,

[0922] a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 17 or a variant thereof, and

[0923] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 18 or a variant thereof.

[0924] 244. The antibody or antigen binding fragment thereof of item 243, wherein the variant of SEQ ID NO: 13 has

[0925] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0926] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except Y or W.

[0927] 245. The antibody or antigen binding fragment thereof of item 243 or 244, wherein the variant of SEQ ID NO: 13 has

[0928] (i) at most two, in particular one, amino acid substitutions at any one of positions 1, 2, 4 and / or 5, and / or

[0929] (ii) an amino acid substitution of A at position 3 any amino acid, in particular canonical amino acid, except R, I, L, E, F, K, Y or W. 246. The antibody or antigen binding fragment thereof of any one of items 243 to 245, wherein the variant of SEQ ID NO: 14 has

[0930] (i) at most four, in particular two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 16;

[0931] (ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

[0932] 247. The antibody or antigen binding fragment thereof of item 246, wherein the variant of SEQ ID NO: 14 has

[0933] (i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 2, 5 and / or any one of positions 7 to 16.

[0934] 248. The antibody or antigen binding fragment thereof of any one of items 243 to 247, wherein the variant of SEQ ID NO: 15 has

[0935] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or

[0936] (ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

[0937] 249. The antibody or antigen binding fragment thereof of any one of items 243 to 248, wherein the variant of SEQ ID NO: 15 has

[0938] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 6, 8, 9, 15 and / or any one of positions 16 to 20, and / or

[0939] (ii) an amino acid substitution of D at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except I, D, L, R or W, an amino acid substitution of G at position 7 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of G at position 10 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except W or P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except S, D, N, Y, E, L, G, C or P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except H, L, P, F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except R, H, Q, F, L, Y, K or W.

[0940] 250. The antibody or antigen binding fragment thereof of any one of items 243 to 249, wherein the variant of SEQ ID NO: 16 has

[0941] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or

[0942] (ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

[0943] 251. The antibody or antigen binding fragment thereof of item 250, wherein the variant of SEQ ID NO: 16 has

[0944] (i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 10.

[0945] 252. The antibody or antigen binding fragment thereof of any one of items 243 to 251, wherein the variant of SEQ ID NO: 18 has

[0946] (i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or

[0947] (ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

[0948] 253. The antibody or antigen binding fragment thereof of any one of items 243 to 252, wherein the variant of SEQ ID NO: 18 has

[0949] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 10 and / or 12, and / or

[0950] (ii) an amino acid substitution of A at position 1 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except K, Q, V, R, W, T, Y, F, H, M, L or I, and / or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except F, Y, L or W.

[0951] 254. The antibody or antigen binding fragment thereof of item 252 or 253, wherein the variant of SEQ ID NO: 18 has

[0952] (i) at most two, in particular one, amino acid substitutions at positions 2, any one of positions 4 to 7, positions 9, 10 and / or 12.

[0953] 255. The antibody or antigen binding fragment thereof of any one of items 252 to 254, wherein the variant of SEQ ID NO: 18 has

[0954] (i) at most two, in particular one, amino acid substitutions at positions 6, 10 and / or 12.

[0955] 256. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: 31 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 32 or variant thereof.

[0956] 257. The antibody or antigen binding fragment thereof of item 256, wherein the variant of SEQ ID NO: 31 is having at least 85 % sequence identity.

[0957] 258. The antibody or antigen binding fragment thereof of item 256 or 257, wherein the variant of SEQ ID NO: 31 is having at least 90 % sequence identity.

[0958] 259. The antibody or antigen binding fragment thereof of any one of items 256 to 258, wherein the variant of SEQ ID NO: 31 is having at least 95 % sequence identity.

[0959] 260. The antibody or antigen binding fragment thereof of any one of items 256 to 259, wherein the variant of SEQ ID NO: 32 is having at least 85 % sequence identity.

[0960] 261. The antibody or antigen binding fragment thereof of any one of items 256 to 260, wherein the variant of SEQ ID NO: 32 is having at least 90 % sequence identity.

[0961] 262. The antibody or antigen binding fragment thereof of any one of items 256 to 261, wherein the variant of SEQ ID NO: 32 is having at least 95 % sequence identity.

[0962] 263. The antibody or antigen binding fragment thereof of any one of items 256 to 262, wherein the heavy chain variable domain comprises a CDR-H1 comprising the amino acid sequence as shown in SEQ ID NO: 19 or a variant thereof,

[0963] a CDR-H2 comprising the amino acid sequence as shown in SEQ ID NO: 20 or a variant thereof, and

[0964] a CDR-H3 comprising the amino acid sequence as shown in SEQ ID NO: 21 or a variant thereof; and / or

[0965] the light chain variable domain comprises

[0966] a CDR-L1 comprising the amino acid sequence as shown in SEQ ID NO: 22 or a variant thereof,

[0967] a CDR-L2 comprising the amino acid sequence as shown in SEQ ID NO: 23 or a variant thereof, and

[0968] a CDR-L3 comprising the amino acid sequence as shown in SEQ ID NO: 24 or a variant thereof.

[0969] 264. The antibody or antigen binding fragment thereof of any one of items 1 to 263, wherein the variant of the sequence comprised in the antibody or antigen binding fragment thereof is a variant with at most four, in particular at most three, in particular at most two amino acid substitutions, in particular one amino acid substitution.

[0970] 265. The antibody or antigen binding fragment thereof of any one of items 1 to 264, wherein the antibody or antigen binding fragment thereof binds to an epitope comprising the sugar moiety and the amino acid residues Val1, His2 and Thr4 as shown in SEQ ID NO: 33 or a variant thereof.

[0971] 266. The antibody or antigen binding fragment thereof of item 265, wherein the antibody or antigen binding fragment thereof binds to an epitope comprising the sugar moiety and the amino acid residues Val1, His2 and Thr4 as shown in SEQ ID NO: 33.

[0972] 267. The antibody or antigen binding fragment thereof of any one of items 1 to 264, wherein the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions.

[0973] 268. The antibody or antigen binding fragment thereof of item 267, wherein the variant is a variant with one amino acid substitution. -110-269. The antibody or antigen binding fragment thereof of any one of items 1 to 264, wherein the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the sequence as shown in SEQ ID NO: 33.

[0974] 270. The antibody or antigen binding fragment thereof of any one of items 1 to 264, wherein the antibody or antigen binding fragment thereof binds to an antigen comprising the sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions.

[0975] 271. The antibody or antigen binding fragment thereof of item 270, wherein the variant is a variant with one amino acid substitution.

[0976] 272. The antibody or antigen binding fragment thereof of any one of items 1 to 264, wherein the antibody or antigen binding fragment thereof binds to an antigen comprising the sequence as shown in SEQ ID NO: 33.

[0977] 273. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc), wherein the antibody or antigen binding fragment thereof binds to an epitope comprising the sugar moiety, the amino acid residues Val1, His2 and Thr4 as shown in SEQ ID NO: 33 or a variant thereof.

[0978] 274. The antibody or antigen binding fragment thereof of item 273, wherein the antibody or antigen binding fragment thereof binds to an epitope comprising the sugar moiety and the amino acid residues Val1, His2 and Thr4 as shown in SEQ ID NO: 33.

[0979] 275. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc), wherein the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions.

[0980] 276. The antibody or antigen binding fragment thereof of item 275, wherein the variant is a variant with one amino acid substitution.

[0981] 277. The antibody or antigen binding fragment thereof of item 275, wherein the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33. 278. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc), wherein the antibody or antigen binding fragment thereof binds to an antigen comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions.

[0982] 279. The antibody or antigen binding fragment thereof of item 278, wherein the variant is a variant with one amino acid substitution.

[0983] 280. The antibody or antigen binding fragment thereof of item 278, wherein the antibody or antigen binding fragment thereof binds to an antigen comprising or consisting of the amino acid sequence as shown in SEQ ID NO: 33.

[0984] 281. The antibody or antigen binding fragment thereof of any one of items 1 to 280, wherein the variant of SEQ ID NO: 33 is a variant, wherein the one or two, in particular one, of the amino acid substitutions is one of the following amino acid substitutions L3V, L3P, T4P or T4N.

[0985] 282. The antibody or antigen binding fragment thereof of any one of items 1 to 281, wherein all amino acid substitutions of the antibody or antigen binding fragment thereof are conservative amino acid substitutions.

[0986] 283. The antibody or antigen binding fragment thereof of item 282, wherein each of said conservative amino acid substitutions is the substitution of an amino acid with another amino acid selected from its same group, wherein the groups of amino acids are

[0987] a) the nonpolar, hydrophobic amino acids consisting of Gly, Ala, Val, Leu, Ile, Phe, Try, Trp, and Met;

[0988] b) the polar, neutral amino acids consisting of Ser, Thr, Asn, and Gln;

[0989] c) the positively charged, basic amino acids consisting of Arg, Lys, and His, and d) the negatively charged, amino acids consisting of Asp and Glu;

[0990] wherein if Cys is to be conservatively substituted, it is substituted with Ser or Ala, and wherein if Pro is to be conservatively substituted it is substituted with Ala.

[0991] 284. The antibody or antigen binding fragment thereof of item 282 or 283, wherein each of said conservative amino acid substitutions is a highly conservative amino acid substitution selected from

[0992] a) substitution of Ala with Val, Leu, Ile or Gly; b) substitution of Arg with Lys;

[0993] c) substitution of Asn with Gin;

[0994] d) substitution of Asp with Glu;

[0995] e) substitution of Cys with Ser;

[0996] f) substitution of Gin with Asn;

[0997] g) substitution of Glu with Asp;

[0998] h) substitution of Gly with Ala;

[0999] i) substitution of His with Arg;

[1000] j) substitution of Ile with Leu, Val or Ala;

[1001] k) substitution of Leu with Ile, Val or Ala;

[1002] l) substitution of Lys with Arg;

[1003] m) substitution of Met with Leu, Ile or Val;

[1004] n) substitution of Phe with Try or Trp;

[1005] o) substitution of Pro with Ala;

[1006] p) substitution of Ser with Thr;

[1007] q) substitution of Thr with Ser;

[1008] r) substitution of Trp with Phe or Tyr;

[1009] s) substitution of Tyr with Phe or Trp; and

[1010] t) substitution of Val with Leu, Ile or Ala.

[1011] 285. The antibody or antigen binding fragment thereof of any one of items 1 to 284, wherein the antibody is a monoclonal antibody.

[1012] 286. The antibody or antigen binding fragment thereof of any one of items 1 to 284, wherein the antigen binding fragment is a Fab-fragment, a F(ab')2-fragment, a Fab'-fragment, a single-chain variable fragment (scFv), di-scFv, single-domain antibody, or a protein comprising (VL-CL)nand (VH-CHl)n, wherein n is an integer of 2 to 12 and wherein each (VL-CL) or each (VH-CH1) is connected to the next (VL-CL) or next (VH-CH1) via a peptide linker.

[1013] 287. The antibody or antigen binding fragment thereof of any one of items 1 to 286, wherein the antibody or antigen binding fragment thereof is a recombinant antibody or antigen binding fragment thereof. 288. The antibody or antigen binding fragment thereof of any one of items 1 to 287, wherein the antibody or antigen binding fragment thereof binds to HbAlc with a dissociation constant, KD, of 20 nM or lower, preferably wherein the KD is measured at 37 °C.

[1014] 289. The antibody or antigen binding fragment thereof of any one of items 1 to 288, wherein the antibody or antigen binding fragment thereof binds to HbAlc with a dissociation constant, KD, of 0.3 nM to 20 nM, preferably wherein the KD is measured at 37 °C.

[1015] 290. The antibody or antigen binding fragment thereof of any one of items 1 to 289, wherein the antibody or antigen binding fragment thereof binds to HbAlc with an association rate constant (ka) of 25000 M^s’1or higher.

[1016] 291. The antibody or antigen binding fragment thereof of any one of items 1 to 290, wherein the antibody or antigen binding fragment thereof binds to HbAlc with an association rate constant (ka) of 25000 M^s’1to 100000 M’.

[1017] 292. The antibody or antigen binding fragment thereof of any one of items 1 to 291, wherein the antibody or antigen binding fragment thereof binds to HbAlc with an dissociation rate constant (kd) of 0.00024 s-1or lower.

[1018] 293. The antibody or antigen binding fragment thereof of any one of items 1 to 292, wherein the antibody or antigen binding fragment thereof binds to HbAlc with an dissociation rate constant kd) of 0.00024 s’1to 0.001 s’1.

[1019] 294. The antibody or antigen binding fragment thereof of any one of items 1 to 293, wherein the antibody or antigen binding fragment thereof binds to HbAlc with a t / 2 diss of 12 min or more.

[1020] 295. The antibody or antigen binding fragment thereof of any one of items 1 to 294, wherein the antibody or antigen binding fragment thereof binds to HbAlc with a t / 2 diss of 12 min to 46 min.

[1021] 296. The antibody or antigen binding fragment thereof of any one of items 1 to 295, wherein when the antibody or antigen binding fragment thereof is used in a turbidimetric inhibition immunoassay for determining the levels of HbAlc using a sample comprising a normal physiological concentration of HbAlc, the immunoassay has a coefficient of variation of 1.5 % or lower. 297. The antibody or antigen binding fragment thereof of item 296, wherein when the antibody or antigen binding fragment thereof is used in a turbidimetric inhibition immunoassay for determining the levels of HbAlc using a sample comprising a normal physiological concentration of HbAlc, the immunoassay has a coefficient of variation of 0.8 % or lower.

[1022] 298. The antibody or antigen binding fragment thereof of item 296 or 297, wherein the sample is comprising HbAlc at a concentration lower than 6.0 %, wherein the percentage is the amount of HbAlc compared to the total amount of hemoglobin as measured by turbidimetric inhibition immunoassay.

[1023] 299. The antibody or antigen binding fragment thereof of any one of items 1 to 298, wherein when the antibody or antigen binding fragment thereof is used in a turbidimetric inhibition immunoassay for determining the levels of HbAlc using a sample comprising an abnormal physiological concentration of HbAlc, the immunoassay has a coefficient of variation of 1.5 % or lower.

[1024] 300. The antibody or antigen binding fragment thereof of item 299, wherein the sample is comprising HbAlc at a concentration higher than or equal to 6.0 %, wherein the percentage is the amount of HbAlc compared to the total amount of haemoglobin as measured by turbidimetric inhibition immunoassay.

[1025] 301. The antibody or antigen binding fragment thereof of any one of items 1 to 300, wherein the antibody or antigen binding fragment thereof binds to glycated haemoglobin subunit beta.

[1026] 302. The antibody or antigen binding fragment thereof of item 301, wherein the antibody or antigen binding fragment thereof binds to an antigen comprising or consisting of SEQ ID NO: 41 or a variant thereof with one amino acid substitution.

[1027] 303. The antibody or antigen binding fragment thereof of item 302, wherein the variant is L3V, L3P, T4P or T4N.

[1028] 304. The antibody or antigen binding fragment thereof of any one of items 1 to 303, wherein the antibody or antigen binding fragment thereof does not bind to non-glycated hemoglobin subunit beta. 305. The antibody or antigen binding fragment thereof of any one of items 1 to 304, wherein the antibody or antigen binding fragment thereof does not bind to hemoglobin AO (HbAO).

[1029] 306. The antibody or antigen binding fragment thereof of item 305, wherein the antibody or antigen binding fragment thereof does not bind to hemoglobin A0 (HbAO) when tested with ELISA, in particular binding HbAO with an OD of less than 0.1, wherein HbAO is immobilized in a well of a standard 96-well plate with 100 pl of 31.25 ng / ml HbAO and to which well, after removal of the previous solution, are 30 µl of B-cell culture supernatant added.

[1030] 307. A nucleic acid coding for at least a part of an antibody or antigen binding fragment thereof of any one of items 1 to 306.

[1031] 308. The nucleic acid of item 307, wherein the nucleic acid is DNA or RNA.

[1032] 309. The nucleic acid of item 307, wherein the part of an antibody or antigen binding fragment thereof is at least one CDR, at least one light chain variable domain or at least one heavy chain variable domain.

[1033] 310. An expression construct comprising a nucleic acid of any one of items 307 to 309.

[1034] 311. The expression construct of item 310 comprising a promoter, a terminator, a 5' UTR and / or a 3' UTR.

[1035] 312. The expression construct of item 311, wherein the promoter is an inducible promoter.

[1036] 313. The expression construct of item 311, wherein the promoter is a constitutive promoter.

[1037] 314. A vector comprising an expression construct of any one of items 310 to 313.

[1038] 315. The vector of item 314 comprising at least one selection marker and / or at least one origin of replication.

[1039] 316. The vector of item 314 or 315, wherein the vector is a viral vector, artificial chromosome or a plasmid. 317. A cell comprising an antibody or antigen binding fragment thereof of any one of items 1 to 306, a nucleic acid of any one of items 307 to 309, an expression construct of any one of items 310 to 313 or a vector of any one of items 314 to 316.

[1040] 318. The cell of item 317, wherein the cell is a eukaryotic cell, in particular a human cell.

[1041] 319. A composition comprising an antibody or antigen binding fragment thereof of any one of items 1 to 306 and an acceptable carrier.

[1042] 320. A diagnostic composition comprising an antibody or antigen binding fragment thereof of any one of items 1 to 306 and an acceptable carrier.

[1043] 321. A kit comprising an antibody or antigen binding fragment thereof of any one of items 1 to 306, a composition of item 319 or a diagnostic composition of item 320.

[1044] 322. Use of an antibody or antigen binding fragment thereof of any one of items 1 to 306, a composition of item 320 or a diagnostic composition of item 321 for determining the amount or concentration of HbAlc in a sample.

[1045] 323. The use of item 322, wherein the sample is a biological fluid sample.

[1046] 324. The use of item 323, wherein the biological fluid sample is whole blood, in particular capillary or venous blood, serum, blood plasma, saliva or interstitial fluid.

[1047] 325. The use of item 323, wherein the antibody is for use in an immunoassay, in particular an competitive immunoassay.

[1048] 326. Method of determining the levels of HbAlc in a sample comprising a step of bringing the sample into contact with an antibody or antigen binding fragment thereof of any one of items 1 to 306.

[1049] 327. Method of determining the ratio of HbAlc in a sample comprising the following steps:

[1050] (a) adding to the sample an antibody or antigen binding fragment thereof of any one of items 1 to 306,

[1051] (b) measuring with a photometer the absorbance of the sample to determine the total amount of haemoglobin in the sample, (c) adding to the sample a polyhapten comprising at least one binding site for the antibody or antigen binding fragment thereof of any one of items 1 to 306, and

[1052] (d) measuring with a photometer the absorbance of the sample to determine the amount of HbAlc in the sample.

[1053] 328. The method of determining the levels of HbAlc in a sample of item 326 or 327, wherein the absorbance in step (b) is measured at a wavelength of about 500 to about 650 nm, in particular 546 nm.

[1054] 329. The method of determining the levels of HbAlc in a sample of any one of items 326 to 328, wherein the absorbance in step (d) is measured at a wavelength of about 340 to about 600 nm, in particular 340 nm.

[1055] 330. The method of determining the levels of HbAlc in a sample of any one of items 326 to 329, wherein the absorbance measured in step (d) is inversely proportional to the concentration of HbAlc in the sample.

[1056] 331. The method of determining the levels of HbAlc in a sample of any one of items 326 to 330, wherein the sample is whole blood, which has been hemolysed before adding the antibody in step (a).

[1057] 332. The method of determining the levels of HbAlc in a sample of any one of items 326 to 331, wherein the antibody has been added in step (a) to a final concentration of about 0.025 to 10 mg / ml.

[1058] 333. The method of determining the levels of HbAlc in a sample of any one of items 326 to 332, wherein the polyhapten has been added in step (c) to a final concentration of about 2.5 to 50 μg / ml.

[1059] 334. The method of determining the levels of HbAlc in a sample of any one of items 326 to 333, wherein the method is a turbidimetric inhibition immunoassay.

[1060] 335. The method of determining the levels of HbAlc in a sample of any one of items 326 to 334, wherein the method has a coefficient of variation of 1.5 % or lower, when using a sample comprising a normal physiological concentration of HbAlc. 336. The method of determining the levels of HbAlc in a sample of item 335, wherein the method has a coefficient of variation of 0.8 % or lower, when using a sample comprising a normal physiological concentration of HbAlc.

[1061] 337. The method of determining the levels of HbAlc in a sample of item 335 or 336, wherein the sample is comprising HbAlc at a concentration lower than 6.0 %, wherein the percentage is the amount of HbAlc compared to the total amount of hemoglobin.

[1062] 338. The method of determining the levels of HbAlc in a sample of any one of items 326 to 337, wherein the method has a coefficient of variation of 1.5 % or lower, when using a sample comprising an abnormal physiological concentration of HbAlc.

[1063] 339. The method of determining the levels of HbAlc in a sample of item 338, wherein the sample is comprising HbAlc at a concentration higher than or equal to 6.0 %, wherein the percentage is the amount of HbAlc compared to the total amount of hemoglobin. Sequence Listing

[1064] Name Amino Acids SEQ ID

[1065] CDR-H1 54E11 / 55C4 SYAMS SEQ ID NO: 1

[1066] YIWX1X2X3X4TYX5X6X7YASWAKG, wherein Xi is K or S,

[1067] CDR-H2 54E11 / 55C4 X2 is T or G, X3 is R or D, X4 is N or R, X5 is S or not present, SEQ ID NO: 2

[1068] Xe is T or not present, X7 is Y or not present

[1069] CDR-H3 54E11 / 55C4 DYPFYGGX8VGVVDAMSRLDL, wherein X8is D or V SEQ ID NO: 3

[1070] CDR-L1 54E11 / 55C4 QSSQSVYX9NNWLA, wherein X9is N or M SEQ ID NO: 4

[1071] CDR-L2 54E11 / 55C4 YAX10TLAS, wherein X10is S or A SEQ ID NO: 5

[1072] CDR-L3 54E11 / 55C4 AGAYSX11PSGDNG, wherein X11is S or T SEQ ID NO: 6

[1073] CDR-H1 54E11 SYAMS SEQ ID NO: 7

[1074] CDR-H254E11 YI WKTRNT YS T YYAS WAKG SEQ ID NO: 8

[1075] CDR-H3 54E11 DYPFYGGDVGVVDAMSRLDL SEQ ID NO: 9 Name Amino Acids SEQ ID CDR-L1 54E11 QSSQSVYNNNWLA SEQ ID NO: 10 CDR-L2 54E11 YASTLAS SEQ ID NO: 11 CDR-L3 54E11 AGAYSSPSGDNG SEQ ID NO: 12 CDR-H1 55C4 SYAMS SEQ ID NO: 13 CDR-H2 55C4 YIWSGDRTYYASWAKG SEQ ID NO: 14 CDR-H3 55C4 DYPFYGGVVGVVDAMSRLDL SEQ ID NO: 15 CDR-L1 55C4 QSSQSVYMNNWLA SEQ ID NO: 16 CDR-L2 55C4 YAATLAS SEQ ID NO: 17 CDR-L3 55C4 AGAYSTPSGDNG SEQ ID NO: 18 CDR-H1 65H2 GSQSIW SEQ ID NO: 19 CDR-H2 65H2 WIHAGSPGYTGYASWAKG SEQ ID NO: 20 Name Amino Acids SEQ ID

[1076] CDR-H3 65H2 AYPYGGFDI SEQ ID NO: 21

[1077] CDR-L1 65H2 QASESVYSNNDLS SEQ ID NO: 22

[1078] CDR-L265H2 TASSLAS SEQ ID NO: 23

[1079] CDR-L3 65H2 AGYKSTSSDGFA SEQ ID NO: 24

[1080] QSVEESGGRLVTPGTPLTLTCTX12SGFSLSSYAMSWVRQ APGKGLEWX13GYIWX1X2X3X4TYX5X6X7YASWAKGRFTI SKXI4SSTTVDLKMTSPTTEDTATYFCARDYPFYGGX8VG

[1081] VH 54E11 / 55C4 VVDAMSRLDLWGQGTLVTVSS, wherein Xi is K or S, X2is SEQ ID NO: 25

[1082] T or G, X3 is R or D, X4is N or R, X5 is S or not present, Xe is

[1083] T or not present, X7 is Y or not present, X8is D or V, X12 is A

[1084] or V, X13 is V or I, Xi4is T or S

[1085] AVLTQTPSPVSAAVGGTVTIX15CQSSQSVYX9NNWLAWY QQKPGQPPKLLX16YYAX10TLASGVPSRFX17GSGSGTQFT

[1086] VL 54E11 / 55C4 LTISDVQCX18DAATYYCAGAYSXnPSGDNGFGGGTEVV SEQ ID NO: 26

[1087] VK, wherein X9 is N or M, X10 is S or A, X11 is S or T, X15 is S

[1088] or N, X16 is I or V, X17 is T or K, X18 is D or A Name Amino Acids SEQ ID

[1089] QSVEESGGRLVTPGTPLTLTCTASGFSLSSYAMSWVRQA PGKGLEWVGYIWKTRNTYSTYYASWAKGRFTISKTSSTT VH 54E11 SEQ ID NO: 27

[1090] VDLKMTSPTTEDTATYFCARDYPFYGGDVGVVDAMSRL DLWGQGTLVTVSS

[1091] AVLTQTPSPVSAAVGGTVTISCQSSQSVYNNNWLAWYQ VL 54E11 QKPGQPPKLLIYYASTLASGVPSRFTGSGSGTQFTLTISDV SEQ ID NO: 28

[1092] QCDDAATYYCAGAYSSPSGDNGFGGGTEVVVK

[1093] QSVEESGGRLVTPGTPLTLTCTVSGFSLSSYAMSWVRQA PGKGLEWIGYIWSGDRTYYASWAKGRFTISKSSSTTVDL VH 55C4 SEQ ID NO: 29

[1094] KMTSPTTEDTATYFCARDYPFYGGVVGVVDAMSRLDLW GQGTLVTVSS

[1095] AVLTQTPSPVSAAVGGTVTINCQSSQSVYMNNWLAWYQ VL 55C4 QKPGQPPKLLVYYAATLASGVPSRFKGSGSGTQFTLTISD SEQ ID NO: 30

[1096] VQCADAATYYCAGAYSTPSGDNGFGGGTEVVVK

[1097] QSLEESGGDLVQPGASLRLTCKASGLDFSGSQSIWWVRQ VH 65H2 SEQ ID NO: 31

[1098] APGKGLEWIAWIHAGSPGYTGYASWAKGRFTISKTSSTT Name Amino Acids SEQ ID

[1099] VTLQMTSLTAADTATYLCARAYPYGGFDIWGPGTLVTV SS

[1100] IVMTQTPSSKSVPVGDTVTINCQASESVYSNNDLSWYQQ VL 65H2 KPGQPPKLLIYTASSLASGVPSRFKGSGSGTQFTLTISDVV SEQ ID NO: 32

[1101] CDDAATYYCAGYKSTS SDGFAFGGGTEVVVK

[1102] N-l-deoxyfructosyl-VHLT VHLT, wherein a deoxy-fructosyl is attached to the N-terminus SEQ ID NO: 33

[1103] N-l-deoxyfructosyl-VHLT- VHLTC, wherein a deoxy-fructosyl is attached to the N- SEQ ID NO: 34 KLH terminus and a KLH at the C-terminus

[1104] N-l-deoxyfructosyl-VHLT- VHLTE, wherein a deoxy-fructosyl is attached to the N- SEQ ID NO: 35 GluBiPEG4 terminus and GluBiPEG4 at the C-terminus

[1105] VHLT-GluBiPEG4 VHLTE, wherein GluBiPEG4 at the C-terminus SEQ ID NO: 36

[1106] N-l-deoxyfructosyl-VHVT- VHVTEE, wherein a deoxy-fructosyl is attached to the N- SEQ ID NO: 37 GluBiPEG7 (Kamakura) terminus and GluBiPEG4 at the C-terminus Name Amino Acids SEQ ID

[1107] N-l-deoxyfructosyl-VHPT- VHPTEE, wherein a deoxy-fructosyl is attached to the N- SEQ ID NO: 38 GluBiPEG7 (Jabalpur) terminus and GluBiPEG4 at the C-terminus

[1108] N-l-deoxyfructosyl-VHLP- VHLPEE, wherein a deoxy-fructosyl is attached to the N- SEQ ID NO: 39 GluBiPEG7 (Benin City) terminus and GluBiPEG4 at the C-terminus

[1109] N-l-deoxyfructosyl-VHLN- VHLNEE, wherein a deoxy-fructosyl is attached to the N- SEQ ID NO: 40 GluBiPEG7 (Wuerzburg) terminus and GluBiPEG4 at the C-terminus

[1110] VHLTPEEKSAVTALWGKVNVDEVGGEALGRLLVVYPW TQRFFESFGDLSTPDAVMGNPKVKAHGKKVLGAFSDGL

[1111] hemoglobin subunit beta SEQ ID NO: 41

[1112] AHLDNLKGTFATLSELHCDKLHVDPENFRLLGNVLVCVL AHHFGKEFTPPVQAAYQKVVAGVANALAHKYH Examples

[1113] The following examples are provided to illustrate, but not to limit the presently claimed invention.

[1114] Peptide synthesis

[1115] Peptides were synthesized by means of fluorenylmethyloxycarbonyl (Fmoc) solid phase peptide synthesis as described in EP 0 329 994 Al. β N-1-deoxyfructosyl-Val-His-Leu-Thr (HbAlc(l-4), SEQ ID NO: 33) is used as peptide to ensure that common Hb variants are analytically correctly determined. Biotinylated peptides were synthesized using Glu(biotinyl-PEG) as a building block.

[1116] Table 1: Summary of synthesized peptides

[1117] Name Structure

[1118] HbAlc(l-4)-KLH immunogen (SEQ ID NO: 34)

[1119]

[1120] Table 1: Summary of synthesized peptides

[1121] Name Structure

[1122] HbAlc(l- 4)[Glu(BiPEG)-4]amid (SEQ ID NO: 35)

[1123]

[1124] HbA0(l- 4)[Glu(BiPEG)-4]amid (SEQ ID NO: 36)

[1125]

[1126] Immunogen synthesis

[1127] To a solution of keyhole limpet hemocyanin (KLH) (29,9 mg) in phosphate buffer (45 mL, 20 mM, pH 7.2) 6-Maleimidohexanoic acid N-hydroxysuccinimide ester (6.15 mg; 20 pmol dissolved in 1.5 mL DMSO) was added. The reaction was incubated for 5 hours at room temperature and then dialyzed against phosphate buffer (0.1 M, pH 7.2). Cysteine containing HbAlc(l-4) peptide (2.1 mg; 2.9pmol) was dissolved in DMSO and added to a solution of maleimide-activated KLH (25.6 mg) containing 0.1 M EDTA. The solution was agitated over night at 4 °C and then dialyzed against PBS buffer (pH 7.4) to yield 24 mg HbAlc(l-4)-KLH peptide (Table 1, SEQ ID NO: 34) conjugate. Immunization

[1128] New Zealand White (NZW) rabbits, 12-16 weeks old, were immunized with HbAlc(l-4)-KLH (Table 1, SEQ ID NO: 34). To enhance the immunogenicity of the peptide it was coupled to keyhole limpet hemocyanin (KLH) as a carrier protein. In the first month the animals were immunized weekly. Starting in the second month, the immunization schedule was reduced to once per month. For the first immunization 500 pg HbAlc(l-4)-KLH was dissolved in 0.9 % NaCl and emulsified in 2 ml complete Freund’s Adjuvant (CFA). For all following immunizations, CFA was replaced by 1 mL Incomplete Freund’s Adjuvant (IF A) emulsion.

[1129] Titer analysis

[1130] Titer analysis was performed with an ELISA protocol. Serum titrations were performed using HbAlc(l-4)[Glu(BiPEG)-4]amid as a positive control. Biotinylated HbAlc(l-4)[Glu(BiPEG)-4]amid screening reagent (Table 1, SEQ ID NO: 35) was immobilized on the surface of 96 well streptavidin-coated microtitre plates by incubating 100 µl per well of a 31.25 ng / ml solution for 60 min at room temperature. Subsequent washing was performed using an automated instrument (Biotek) according to manufacturer’s instructions. A small amount of serum from each rabbit (2 - 3 ml per animal) was collected on day 45 and day 105 after the start of the immunization campaign. The serum from each rabbit was diluted 1:300, 1:900, 1:2700, 1:8100, 1:24300, 1:72900, 1:218700 and 1: 656100 with PBS containing 1 %BSA. 100 pl of each dilution was added to the plate previously prepared with the screening peptides and incubated for 60 min at room temperature. Bound antibody was detected with a HRP-labeled F(ab')2 goat anti-rabbit Fey (Dianova) and ABTS substrate solution (Roche). The titer of the analyzed animals was set at 50 % signal decrease of the dilution curve.

[1131] Table 2 Four exemplary titers after immunization with HbAlc-KLH

[1132] Animal Titer day 35 Titer day 165

[1133] 1#F37276 17799 6178 Table 2 Four exemplary titers after immunization with HbAlc-KLH

[1134] Animal Titer day 35 Titer day 165

[1135] 2#F34283 2000 7653

[1136] 3#F43699 18598 3720

[1137] 4#F43641 15719 2017

[1138] As demonstrated by the results of Table 2, the polyclonal sera from immunized animals bound to the HbAlc(l-4)[Glu(BiPEG)-4]amid screening peptide.

[1139] B-cell cloning

[1140] For enrichment of antigen reactive B-cells, 31.25 ng / ml HbAlc(l-4)[Glu(BiPEG)-4]amid (SEQ ID NO: 35) was pre-incubated with the peripheral blood mononuclear cell (PBMC) pool from the immunized animals for 15 min at 4 °C. After a washing step, the antigen-reactive B cells bound to HbAlc(l-4)[Glu(BiPEG)-4]amid were incubated with streptavidin-coated beads (Miltenyi) for 15 min at 4 °C. Sorting of positive B-cells using MACS columns (Miltenyi) and subsequent incubation were performed as described in Seeber et al., PLoS One, 2014 Feb 4, 9(2):e86184, with the only exception that the sorting of positive B cells involved MACS columns (Miltenyi) instead of plate binding. After a 7-day culture of B cells in 96-well plates, supernatant was collected for subsequent ELISA analysis and cells were lysed for cloning.

[1141] Antibody Screening

[1142] Subsequently a Hit-ELISA (i.e. an ELIS As testing the binding to the screening reagents) was used to identify B-cells expressing antibodies having desired binding characteristics, i.e. binding the HbAlc(l-4)[Glu(BiPEG)-4]amid (SEQ ID NO: 35), butnotHbA0(l-4)[Glu(BiPEG)-4]amid (Table 1, SEQ ID NO: 36). HbAlc(l-4)[Glu(BiPEG)-4]amid and HbA0(l-4)[Glu(BiPEG)-4]amid were immobilized on the surface of streptavidin- coated 96-well plates (Nunc) by incubation of 100 µl per well of 31.25 ng / ml solutions for 60 min at room temperature, respectively. The plates were washed and 30 pl of rabbit B-cell culture supernatant was transferred to each well and incubated for 1 h at room temperature. For the detection of antibodies bound to the screening agents, HRP-labelled F(ab')2 goat anti-rabbit Fey (Dianova) and ABTS substrate solution (Roche) were used according to manufacturer’s instructions. 2833 clones were identified that bound to HbAlc(l-4)[Glu(BiPEG)-4]amid, but not to HbA0(l-4)[Glu(BiPEG)-4]amid (out of 26 B cell sorting experiments with in total four immunized rabbits) (Table 3). The V regions of 927 clones were cloned into mammalian expression vectors and subsequently expressed in 2 ml of HEK293 cells (described in Seeber et al., PLoS One, 2014 Feb 4, 9(2):e86184). After one week of expression the supernatants of the transfected HEK293 cells, containing rabbit IgG, were then used for an initial SPRBiacore™ based selection of a subset of antibodies fulfilling performance criteria for detailed kinetic analysis (see FIG. 1 A and FIG 2A) and evaluation in the Tinaquant assay (see Table 5).

[1143] Table 3: ELISA results of four exemplary clones producing antibody characterized by specific binding to the positive screening reagent (HbAlc(l-4)) but not to the negative screening reagent (HbA0(l-4)).

[1144] Clone ID OD HbAlc(l-4) OD HbA0(l-4)

[1145] 55C4 2.912 0.022

[1146] 54E11 2.886 0.025

[1147] 57A11 2.927 0.031

[1148] 54C8 1.371 0.022

[1149] Down streaming process for screening and clone selection

[1150] The down streaming process of monoclonal antibody binding to HbAlc was designed to be suitable for the screening of many different culture supernatants from transiently transfected HEK or CHO cells. The yields of the recombinant IgGs from rabbit were in the range of 10 to 500 mg. In a single step purification the recombinant rabbit IgG from a culture supernatant was captured by a Protein A affinity column (Praesto® Jetted A50, Purolite, prepacked column) at pH 7.4 and eluted from the column under acidic conditions at pH 3.7 (200 mM sodium citrate). The IgG was brought back to neutral pH before the respective monoclonal antibody binding to HbAlc is dialyzed against the storage buffer (50 mM KPO4 / 150 mM KC1 / pH 7.4). A final filtration step (0.2 pm membrane filter) was applied before storage at -20 °C. Each lot of the respective monoclonal antibody binding to HbAlc was analyzed according to the following methods: protein concentration at OD 280 nm, analytical size exclusion chromatography and functional testing in the HbAlc turbidimetric inhibition immunoassay.

[1151] Kinetic screening

[1152] The kinetic screening was performed at 37 °C on a GE Healthcare Biacore™ 4000 or 8K instrument. A Biacore™ CM5 Series S sensor was mounted into the respective instrument and preconditioned according to the manufacturer’s instructions. The system buffer was HBS ET pH 6.2, 10 mM HEPES, 150 mM NaCl, 3 mM EDTA, 0.05 % (w / v) Tween20, when biotinylated peptides were used as screening reagent. The system buffer was supplemented with 1 mg / mL CMD (Carboxymethyldextran, Sigma, Cat. No. 86524) and was used as sample buffer for the preparation of the samples and dilution series.

[1153] A rabbit antibody capture system was immobilized on the sensor surface at 25 °C using HBS-N pH 7.4 containing 10 mM HEPES, 150 mM NaCl as a system buffer. A polyclonal goat anti-rabbit IgG Fc capture antibody GARbFcy (Code-No. 111-005-046; Jackson Immuno Research) was amine coupled using the EDC / NHS-chemistry according to the manufacturer’s instructions. 30 pg / mL goat anti rabbit Fc gamma (GARbFcy) in 10 mM sodium acetate buffer pH 4.5 were preconcentrated in the flow cells and covalently bound to the CMD surface with densities between 11000-14000 RU. Free activated carboxyl groups were saturated with 1 M ethanolamine pH 8.5. When using the 8K instrument, flow cells (Fc) 1 of channels 1-8 served as references and Fc 2 of channels 1-8 were used for the interaction measurements. Using the B4000, Spots 1 and 5 were used for the interaction measurements and spots 2 and 4 served as references. Each rabbit antibody from the primary cell supernatant was diluted 1:2 in the sample buffer and was injected at a flow rate of 10 pL / min for 2 minutes. The rabbit antibody Capture Level (CL) in resonance units (RU) was monitored. To achieve high enough sensitivity, an additional molecular mass load for the biotinylated peptides HbAlc(l-4)[Glu(BiPEG)-4]amid (SEQ ID NO: 35), Roche BMO 28.0920002, “303” 1.2 kDa and HbA0(l-4)[4-(O2Oc)3-Glu(BiPEG)]amid, Roche BMO 35.190039, ”284”, 1.3 kDa was generated by Streptavidin(SA)-grafting. The biotinylated peptides (Table 1), Streptavidin and Amino-PEO-Biotin (Thermo Fisher, Cat. No. 21346) were mixed with a ratio of 1: 10:5 and were incubated for 2 hours at room temperature (RT). This resulted in highly stable complexes HbAlc(l-4)[Glu(BiPEG)-4]amid-SA and HbA0(l-14)[14-Glu(Bi-PEG)]amid-SA with a molecular mass of approx. 61 kDa each. Additionally, the glycated Albumin, Sigma, Cat. No. SLCB1080 was used as screening reagent. Mass loaded peptides and protein were injected as analytes with a single concentration c = 150 nM to the respective surface displayed anti-HbAlc antibodies at 30 pL / min. The association and dissociation phases were monitored for 5 minutes. The rabbit clones were regenerated from the sensor surface by an injection of 10 mM Glycine pH 2.0 at 20 pL / min for 20 seconds, followed by 2 injections of 10 mM Glycine pH 2.25 for 60 seconds. Report points Binding Late (BL), shortly before the end of the analyte injection and Stability Late (SL), shortly before the end of the dissociation phase, were monitored. BL and SL data were used to characterize the antibody / antigen binding stability. Furthermore, the dissociation rate constant kd[s-1] was calculated according to a Langmuir 1:1 model. The antigen / antibody complex stability half-life time (minutes) was calculated according to the formula ln(2) / 60*kd.

[1154] The Molar Ratio, representing the binding stoichiometry, was calculated with the formula:

[1155] MW (antibody) / MW (antigen) * BL (antigen) / CL (antibody)

[1156] Kinetic profile were monitored by the BIAcore™ B4000 Control SW VI.1 and evaluated with the Evaluation SW VI.1 resp. by the BIAcore™ 8K Control-SW V3.0.11.15423 and evaluated by the BIAcore™ Insight Evaluation SW V3.0.11.15423. From a plurality of antibodies, specific antibody kinetics were identified, where the antibodies discriminated between the HbAlc(l-4)-peptide and the negative controls HbA0(l-4) peptides resp. glycated Albumin. Antibodies with a superior kinetic profile for binding the HbAlc(l-4)-peptide were selected: Antibodies with moderate to fast complex formation and different half-life-times were selected as candidates for further characterization. Refer to FIG. 1A for exemplary kinetic profiles for antibodies binding to streptavidin-grafted HbAlc peptide. Mouse-derived prior art antibodies were analyzed analogous using a mouse antibody capture system. A polyclonal rabbit anti-mouse IgG Fc capture antibody RAMFcy (Code-No.315-005-046; Jackson Immuno Research) was amine coupled at 25°C using the EDC / NHS-chemistry according to the manufacturer’s instructions. 30 pg / mL RAMFcy in 10 mM sodium acetate buffer pH 4.5 were preconcentrated in the flow cells and covalently bound to the CMD surface with densities between 11000-14000 RU. Mouse Abs were diluted to 3 nM and captured and analyzed for the target binding. Refer to FIG. IB for kinetic profiles of the analyzed prior art antibodies binding to streptavidin-grafted HbAlc peptide.

[1157] Detailed Kinetic characterization

[1158] Additionally, clones were kinetically analyzed in detail for binding the peptide HbAlc(l-4)[Glu(BiPEG)-4]amid without mass-load at 37 °C. Instead of a single concentration, a series of increasing concentrations of the HbAlc-peptide, c = 11.1 - 270 nM, were analyzed as described for the kinetic screening. Duplicates for concentration 30 nM were analyzed. The association and dissociation rate constants kaand kdwere calculated according to a Langmuir 1: 1 model. Interactions were determined using a BIAcore™ 8K + instrument from GE Healthcare. The system and sample buffer were applied as described, but supplemented with 0.2 mg / mL Bovine Serum Albumin (BSA). Refer to FIG. 2A for the detailed kinetic characterization for clones resp. refer to FIG. 2B for prior art antibodies.

[1159] The kinetic rate constants and the dissociation equilibrium constants KD were determined using a Langmuir 1:1 fitting model according to the BIAcore™Insight Evaluation SW V3.0.11.15423. Secondly, the Langmuir 1:1 fitting Scrubber-SW V2.0c was applied. The 37 °C kinetics of clone 55C4 (see SEQ ID NO: 30 & SEQ ID NO: 29 for the light and heavy chains of the variable domain) were determined with ka3.6E+05 ± 0.1 % M’1, kd 2.6E-04 ± 0.1 % s’1, t / 2 diss = 44 ± 0.1 %; The affinity was KD = 718 ± 0.1 % pM, see table 4. Determined kinetics constants for antibodies 55C4, 54E11, 65H2* and mouse antibodies from Boehringer Mannheim M-810, M-3.230.140 and M-3.51.56 binding to the HbAlc-peptide are stated in table 4. Table 4 SPR-1 Kinetic constants and affinities for a subset of clones binding HbAlc-peptide

[1160] mAb kakdt / 2dissKDRmaxCL MR [M-1s-1] [s-1] [min] [M] [RU] [RU]

[1161] 55C4 3.6E+05 2.6E-04 44 7.2E-10 16.6 967 2.2 ± 0.1 % ± 0.1 % ± 0.1 %

[1162] 54E11 1.8E +05 2.6E-03 4 1.4E-8 5.1 1097 0.6 ± 0.7 % ± 0.8 % ± 1.0 %

[1163] 65H2 * 1.8E +05 5.4E-03 2 3.0E-8 11 1028 1.4 ± 1.0 % ± 0.7 % ± 1.3 %

[1164] M-8A10 2.5E +05 2.0E-04 6 8.0E-9 6.6 807 0.9 ± 0.2 % ± 0.1 % ± 0.3 %

[1165] M-3.230.140 2.2E +05 8.2E-04 14 3.7E-9 6.3 869 0.9 ± 0.1 % ± 0.2 % ± 0.2 %

[1166] M-3.51.56 5.5E +05 6.5E-04 18 1.2E-9 5.0 746 0.8

[1167]

[1168] ± 0.2 % ± 0.6 % ± 0.5 %

[1169] Antibodies marked with a star *) show complex binding behavior, not obeying Langmuir law. Therefore, the stated constants represent apparent values.

[1170] Functional antibody characterization in competitive test setting

[1171] To evaluate the functional compatibility of the preselected monoclonal antibodies with the competitive two component turbidimetric inhibition immunoassay (TINA) for the quantitative (quant) in vitro determination of hemoglobin Ale (HbAlc), the commercially available Tina-quant® Hemoglobin Ale Gen.3 test (Mat. Nr. 07559674) was used on the cobas® c 513 analyzer as described in the instructions of use as well as the respective operating manual for analyzer-specific test instructions. The Tina-quant® hemoglobin Ale Gen.3 test principal has been described in detail in EP 0329994 Al, EP 0559 164 Al and EP 0598329 Al.

[1172] To facilitate monitoring and diagnosis using the competitive test format, stable binding to the denatured native HbAlc protein is essential prior to the addition of the agglutinator Polyhapten. Hence, the clones were analyzed based on the criteria of within run precision using the coefficient of variation (CV) of < 2.5 % for the measuring range of 4.20 % to 20.1 % HbAlc (NGSP) in comparison to the Tina-quant® Hemoglobin Ale Gen.3 assay using a sheep-derived polyclonal antibody (Table 5). The coefficient of variation (CV) is the determined standard deviation divided by the mean: CV = standard deviation / sample mean (n=21) x 100

[1173] The R1 antibody reagent was prepared in accordance to EP 0 598 329 Al replacing the polyclonal antibody (pAb) with the monoclonal antibody at a concentration of 280 pg / ml. The agglutinator Polyhapten containing R3 reagent was directly used from the commercially available Tina-quant® Hemoglobin A1CX Gen.3 test kit (A1CX3). After transfer of the R1 and R3 reagent to the cobas® kit container, the kits were loaded to the respective analyzers according to the instructions of use and the operating manual using the official Tina-quant® Hemoglobin Ale Gen.3 application settings. Testing was performed using the C.f.a.s. HbAlc calibrator (Mat. Nr.: 04528417, Roche) and the liquid ready to use controls, PreciControl HbAlc norm (PCN; Mat. Nr.: 05479207, Roche) and PreciControl HbAlc path (PCP; Mat. Nr.: 05912504, Roche).

[1174] Based on the acquired precision data, the 54E11 (see SEQ ID NO: 28 & SEQ ID NO: 27 for the light and heavy chains of the variable domain) and 55C4 (see SEQ ID NO: 30 & SEQ ID NO: 29 for the light and heavy chains of the variable domain) monoclonal antibodies, but in particular the 55C4 monoclonal antibody, appear to have advantages over the 65H2 (see SEQ ID NO: 32 & SEQ ID NO: 31 for the light and heavy chains of the variable domain), M-3.51.56, M-3.230.140 and M-8A10 antibodies in a competitive test setting with a comparable performance to the Tina-quant Hemoglobin Ale Gen.3 test (Table 5).

[1175] Table 5: Within-run precision analysis of representative monoclonal antibodies in comparison to the A1CX3 test on the cobas® c 513 system

[1176] M- A1CX3 M- M- 65 54E

[1177] mAh 8A1 55C4 (pAb) 3.51.56 3.230.140 H2 11

[1178] 0

[1179] % CV of PCN 5.00 5.0 1.30 0.690 0.500 % 3.90 % 2.90 %

[1180] (n=21) % 1 % 0 % %

[1181] % CV PCP 4.90 1.7 0.70 0.957 0.600 % 4.00 % 3.20 %

[1182] (n=21) % 9 % 0 % % Kinetic characterization for selected antibody Fab fragment 55C4 binding to native protein HbAlc

[1183] To measure the affinity for the interaction of antibody 55C4 with native protein HbAlc, Ident-Nr: 03019063 Roche in. -house Lot 30386800 an inverse assay format was used.

[1184] The antibody Fab Fragment 55C4 was used as analyte in solution to guarantee the monovalent binding of the antibody to the surface-displayed native protein HbAlc. The native HbAlc protein was immobilized on a Cl sensor surface at 25 °C. The measurements were performed using a BIAcore™ T200 instrument from GE Healthcare. The 2-Dimensional Sensor Cl was preconditioned according to the manufacturer’s instructions. HbAlc protein was amine coupled using the EDC / NHS -chemistry according to the manufacturer’s instructions. BSA was used as a control on the reference surface. 25 pg / pL BSA resp. HbAlc in 10 mM sodium acetate buffer pH 4.5 resp. 5.0 were preconcentrated to the reference flow cell respectively the measuring flow cell and covalently bound to the sensor surface with densities of 90-180 RU (BSA) resp.

[1185] 190-375 (HbAlc). Free activated carboxyl groups were saturated with 1 M ethanolamine pH 8.5.

[1186] Flow cell 1 with immobilized BSA served as reference and Flow cell 2 served as measuring cell with immobilized HbAlc protein. The system and sample buffer were used as stated for the kinetic screening. A series with increasing concentrations of the antibody Fab fragment 55C4, c = 3.3-270 nM, dilution factor 3, was injected for 3 minutes, the dissociation was monitored for 5 minutes at a flow rate 30 pL / min at 37 °C. Duplicates were measured for concentration 30 nM to demonstrate reproducibility. After each step, regeneration was performed using 2 injections with 60 seconds 10 mM Glycine pH 2.0. The data evaluation was performed as stated under detailed kinetic characterization.

[1187] The obtained kinetic profile for antibody 55C4 Fab fragment binding to surface displayed native protein HbAlc is shown in FIG. 3. The obtained kinetic constants and the resulting affinity are summarized in Table 6.

[1188] Table 6: Kinetic constants and affinity for clone 55C4 Fab Fragment binding to surface- displayed native protein HbAlc

[1189] ,,, t / 2diss Rmax M mAb ka[M-1s-1] kd[s-1] KD[M]

[1190] [mm] [RU] R Table 6: Kinetic constants and affinity for clone 55C4 Fab Fragment binding to surface- displayed native protein HbAlc

[1191] 55C4 5.0E+04 ± 0.1 4.9E-04 ± 0.4 9.89E-9 ± 0.

[1192] 24

[1193]

[1194] Fab % % 0.4 % 6

[1195] Crystallization of Fab 55C4 with and without HbAlc peptide

[1196] Monoclonal antibody 55C4 IgG was dialyzed against fragmentation conditions (50 mM KPO4, 150 mM KC1, 2 mM EDTA pH 7.4). The fragmentation process started with the addition of 25 mM Cysteine and 17 mU of Papain per mg of IgG. The cleavage of the IgG into Fab and Fc fragments was completed after 155 min. The reaction was stopped by adding 30 mM iodacetamide. At the end of the fragmentation process, > 95 % of the IgG was converted into Fab fragments. The remaining IgG and the resulting Fc fragments were removed by affinity chromatography using a Protein A affinity column (Praesto® Jetted A50, Purolite, prepacked column). The Fab fragments were further purified using two immunoaffinity columns specifically for rabbit-Fc and Papain. The final purity of the Fab fragments was analyzed with SDS-PAGE and the homogeneity of the Fab fragment preparation was shown by ESI-MS data. A solution containing the 55C4 Fab fragment was concentrated to 15 mg / ml and subject to crystallization screening. Crystallization droplets were set up at 21 °C by mixing 100 nl of protein solution with 100 nl of reservoir solution (1:1 ratio), or 140 nl of protein solution with 60 nl of reservoir solution (7:3 ratio) in a vapour diffusion sitting drop experiment. Crystals appeared in several conditions containing polyethylene glycol (PEG) as a precipitating agent.

[1197] Crystals used for structure determination of the Fab without the HbAlc peptide (apo) appeared within two days and grew to full size within three days in a condition containing the following precipitant solution: 30 mM Sodium Nitrate, 30 mM Sodium Bromide, 30 mM Sodium Iodide, 12.5 % w / v 2-Methyl-2,4-pentanediol, 12.5 % w / v PEG 1000, 12.5 % w / v PEG 3350, 100 mM Sodium HEPES-MOPS pH 7.5. Crystals were directly harvested from droplets with a ratio of 7:3 and flash-cooled in liquid N2.

[1198] Crystals used for structure determination of the Fab bound to HbAlc peptide appeared after eight days and grew to full size within thirteen days in a condition containing the following precipitant solution: 30 mM Diethylene glycol, 30 mM Triethylene glycol, 30 mM Tetraethylene glycol, 30 mM Pentaethylene glycol, 12.5 % w / v 2-Methyl-2,4-pentanediol, 12.5 % w / v PEG 1000, 12.5 % w / v PEG 3350, 100 mM Sodium HEPES-MOPS pH 7.5. Crystals from droplets with a ratio of 7:3 were soaked with a solution containing l-7mide HbAlc peptide for 18 hours, resulting in a final concentration of 2 mM of peptide. Soaked crystals were harvested from droplet and flash-cooled in liquid N2.

[1199] Diffraction images were collected with an EIGER2X 16M detector at a temperature of 100 K at the beam line X10SA of the Swiss Light Source and images were processed with the XDS package [Kabsch W. XDS. Acta Cryst. D66, 125-132 (2010)]. For the apo crystal, data from a single crystal were merged to yield a 1.27 A resolution data set in space group P212121 with one molecule in the asymmetric unit (see Table 7). For the HbAlc peptide bound crystal, similarly, data from a single crystal were merged to yield a 1.27 A resolution data set in space group P212121 with one molecule in the asymmetric unit. The structure was determined by molecular replacement with the program PHASER [McCoy AJ, Grosse-Kunstleve RW, Adams PD, Winn MD, Storom LC, Read RJ J. Appl. Cryst. 40, 658-674 (2007)] as a part of the PHENIX suite [Liebschner D. et al. Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Acta Cryst. D75, 861-877 (2019)]. A Fab fragment from PDB-ID 6LDX was split into constant and variable domains and used as search models. The molecular replacement solution model was rebuilt in COOT [Emsley P, Lohkamp B, Scott WG, Cowtan K. Features and development of Coot. Acta Cryst. D66, 486-501 (2010)] and refined with PHENIX Refine.

[1200] Table 7: Data collection and crystallographic refinement statistics for 55C4 with and without HbAlc peptide.

[1201] Data Collection Apo crystal l-7mide HbAlc bound

[1202] Wavelength (A) 1.0 1.0

[1203] Resolution1(A) 63.60 - 1.27 (1.42 - 1.27) 63.65 - 1.27 (1.37-1.27) Spacegroup P 2 1 2 1 2 1 P 2 1 2 1 2 1

[1204] 50.08, 63.64, 121.15, 90.00, 50.10, 63.65, 121.19, 90.00 Unit cell (A, °)

[1205] 90.00, 90.00 90.00, 90.00

[1206] Total reflections 954,010 (45,522) 553,749 (27.884)

[1207] Unique reflections 72,517 (3,627) 83,835 (4,192)

[1208] Multiplicity 13.2 (13.7) 6.6 (6.7)

[1209] Completeness (%) 94.7 (66.3) 95.0 (60.0)

[1210] Mean I / o(I) 15.2 (1.7) 11.1 (1.4)

[1211] Wilson B-factor 15.30 17.5

[1212] R-meas 0.084 (1.670) 0.079 (1.259)

[1213] CCI / 2 0.999 (0.493) 0.999 (0.607)

[1214] Refinement

[1215] Reflections used in

[1216] 72,428 (82) 77,379 (347) refinement

[1217] Reflections used for R- 3579 (4) 4064 (17)

[1218] free

[1219] R-work 14.67 13.75

[1220] R-free 18.57 18.47 Number of non-hydrogen

[1221] 3,729 3,803

[1222] atoms

[1223] macromolecules 3,324 3,317

[1224] Protein residues 438 438

[1225] RMS bonds (A) 0.011 0.011

[1226] RMS angles (°) 1.164 1.649

[1227] Ramachandran favoured

[1228] 97.88 97.97

[1229] (%)

[1230] Ramachandran allowed

[1231] 2.12 2.23

[1232] (%)

[1233] Ramachandran outliers

[1234] 0.00 0.50

[1235] (%)

[1236] Rotamer outliers (%) 0.77 1.34

[1237] Clashscore 1.96 2.23

[1238] Average B-factor (A2) 19.91 17.633

[1239] 1Values in parentheses refer to the highest resolution bins.

[1240] 2Rmeas= Σ|I-| / ΣI where I is intensity.

[1241] 3Rwork= Σ|Fo-<Fc>| / ΣFowhere Fois the observed and Fcis the calculated structure factor amplitude.

[1242] 4Rfree was calculated based on 5 % of the total data omitted during refinement. The structure of Fab 55C4 with and without HbAlc peptide

[1243] To characterize the binding of Fab 55C4 to HbAlc peptide in atomic details the crystal structure of the Fab in the apo form and in the l-7amide HbAlc peptide bound form was determined. The overall conformation of the Fab is highly similar in both the apo and the ligand bound form with an RMSD of 0.11 A. No significant differences are observed in side chains of the CDR-loops, suggesting a fast association rate constant for ligand binding. Since only the first four residues of the peptide were used for immunization these will be the focus of the structural analysis. The remaining three residues of the crystallized peptide were partially visible in the crystal structure and built.

[1244] The antibody paratope is characterized by an unusually long heavy chain CDR3 loop, which creates a large surface area of interaction with the epitope, particularly with the sugar moiety. The heavy chain CDR3 loop occupies an area between the heavy and the light chain constant regions and pushes the two regions slightly further apart than expected for other canonical rabbit antibodies. The sugar moiety (beta-D-fructopyranose) is buried deep in a pocket formed by the heavy and light chain CDR3 loops while the rest of the ligand peptide lies in a groove lined by the heavy chain CDR loops. An analysis with the program PISA Krissinel E and HenrickK. Inference of macromolecular assemblies from crystalline state. J. Mol. Biol. 372, 774— 797 (2007) revealed that a surface area of 663 A2of the ligand is buried by 55C4, which is 65 % of the total ligand surface area. A total of 20 amino acids constitute the paratope, 11 from the heavy chain and 9 from the light chain (see Table 8). However, the interactions between the sugar moiety and the antibody are largely dominated by an intricate hydrogen-bonding network with the hydroxyl groups of the sugar that includes several coordinated water molecules and backbone and side chain atoms of many residues of the paratope. Also present around the sugar are several hydrophobic side chains of the paratope that help to shape the pocket. The peptide forms various hydrophobic and hydrogen-bond interactions. Almost every nitrogen or oxygen atom that makes up the peptide backbone is involved in a hydrogen bond directly with the paratope or via a coordinated water molecule. The side chains of the first two residues of the peptide (see Table 1 and for example SEQ ID NO: 34), VAL1 and HIS2, lie in a part of the groove that is lined by several hydrophobic and aromatic residues. The side chain of the third amino acid, LEU3, however makes no interactions while the hydroxyl group of the last amino acid, THR4, forms a hydrogen bond with heavy chain SER53 (Kabat Numbering). An overview of the ligand binding can be seen in FIG. 4. Overall the interactions between the epitope and the paratope are very extensive and appear highly specific to the sugar moiety and peptide residues 1, 2 and 4.

[1245] Table 8: Summary of Fab 55C4 residue constituents of the paratope, identified with a distance cutoff of 4.5 A.

[1246] Residu

[1247]

[1248] C (Kabat

[1249] Ch D Numb Side Interaction with ain R ering) chain / backbone Interaction type sugar / peptide

[1250] He

[1251] av

[1252] y 2 TYR50 Side chain H-bond Both

[1253] He

[1254] av H-bond &

[1255] y 2 TRP52 Side chain hydrophobic Both

[1256] He

[1257] av

[1258] y 2 SER53 Side chain H-bond Peptide

[1259] He

[1260] av ARG5

[1261] y 2 6 Side chain H-bond Peptide He

[1262] av

[1263] y 2 TYR58 Side chain Hydrophobic Sugar

[1264] He

[1265] av GLY1

[1266] y 3 00D Backbone H-bond Peptide

[1267] He

[1268] av VAL1 H-bond and y 3 00E Both hydrophobic Peptide

[1269] He

[1270] av VAL1

[1271] y 3 OOF Backbone H-bond Peptide

[1272] He

[1273] av ASP 10

[1274] y 3 0G Backbone H-bond Sugar

[1275] He

[1276] av ALAI

[1277] y 3 OOH Side chain Hydrophobic Sugar

[1278] He

[1279] av MET1

[1280] y 3 001 Backbone H2O coordination Sugar

[1281] Li

[1282] ght 1 TYR28 Side chain Hydrophobic Peptide Li

[1283] ght 1 TRP32 Side chain Hydrophobic Peptide

[1284] Li GLY9

[1285] ght 3 0 Backbone H2O coordination Sugar

[1286] Li ALA9

[1287] ght 3 1 Side chain Hydrophobic Both

[1288] Li

[1289] ght 3 TYR92 Backbone H-bond Both

[1290] Li

[1291] ght 3 SER93 Backbone H-bond Peptide

[1292] Li H-bond &

[1293] ght 3 PRO95 Both hydrophobic Sugar

[1294] Li SER95

[1295] ght 3 A Backbone H-bond Sugar

[1296] Li

[1297] ght 3 ASN96 Side chain H-bond Sugar

[1298] Summary:

[1299] • The Fab 55C4 paratope is dominated by an unusually large heavy chain CDR3 loop that forms a large interaction area for the ligand.

[1300] • The conformational similarity between the apo and the ligand bound state of the Fab suggests a fast ligand binding association rate constant.

[1301] • The sugar moiety of the ligand is buried deep in a pocket between the heavy and light chain CDR3 loops and is largely characterised by hydrogen bonds.

[1302] • The interaction between the peptide part of the ligand and the paratope is a mixture of hydrogen bonds and hydrophobic interactions. • The ligand specificity is determined by the sugar moiety and residues 1, 2 and 4 of the peptide.

[1303] • CDR-H1 and CDR-L2 region do not contribute to the binding.

[1304] Crystallization of Fab 65H2 with and without HbAlc peptide:

[1305] Monoclonal antibody 65H2 IgG was dialyzed against fragmentation conditions (50 mM KPO4, 150 mM KC1, 2 mM EDTA pH 7.4). The fragmentation process started with the addition of 25 mM Cysteine and 17 mU of Papain per mg of IgG. The cleavage of the IgG into Fab and Fc fragments was completed after 155 min. The reaction was stopped by adding 30 mM iodacetamide. The exact conditions for fragmentation of IgG with Papain has to be optimized for individual clones. At the end of the fragmentation process, > 95 % of the IgG was converted into Fab fragments. The remaining IgG and the resulting Fcg fragments were removed by affinity chromatography using a Protein A affinity column (Praesto® Jetted A50, Purolite, prepacked column). The Fab fragments were further purified using two immunoaffinity columns specifically for rabbit-Fc and Papain. The final purity of the Fab fragments was analysed with SDS-PAGE and the homogeneity of the Fab fragment preparation was shown by ESI-MS data.

[1306] A solution containing the 65H2 Fab fragment was subject to crystallization screening. The solution contained 20 mM HEPES pH 7.5 and 100 mM NaCl. Crystallization droplets were set up at 21 °C by mixing 100 nl of protein solution with 100 nl of reservoir solution (1:1 ratio), or 140 nl of protein solution with 60 nl of reservoir solution (7:3 ratio) in a vapour diffusion sitting drop experiment. Crystals ap...

Claims

Patent Claims1. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a CDR-H2 as shown in SEQ ID NO: 2 or variant thereof, a CDR-H3 as shown in SEQ ID NO: 3 or variant thereof, a CDR-L1 as shown in SEQ ID NO: 4 or variant thereof, and a CDR-L3 as shown in SEQ ID NO: 6 or variant thereof.

2. The antibody or antigen binding fragment thereof of claim 1, wherein the variant of SEQ ID NO: 2 has(i) at most four, in particular at most two, in particular one, amino acid substitutions to any amino acid at positions 1, 2 and / or any one of positions 4 to 19, and / or(ii) an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except V or T, in particular an amino acid substitution of W at position 3 to any amino acid, in particular canonical amino acid, except N, A, L, M, C, I, G, S, R, K, E, Y, D, P, Q, H, V or T.

3. The antibody or antigen binding fragment thereof of claim 1 or 2, wherein the variant of SEQ ID NO: 3 has(i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 6, 7, 8, 9, 10, 15 and / or any one of positions 16 to 20, and / or(ii) an amino acid substitution of P at position 3 to any amino acid, in particular canonical amino acid, except W, an amino acid substitution of V at position 11 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of V at position 12 to any amino acid, in particular canonical amino acid, except P, an amino acid substitution of D at position 13 to any amino acid, in particular canonical amino acid, except F, R, W or Y, and / or an amino acid substitution of A at position 14 to any amino acid, in particular canonical amino acid, except W.

4. The antibody or antigen binding fragment thereof of any one of claims 1 to 3, wherein the variant of SEQ ID NO: 4 has(i) at most two, in particular one, amino acid substitutions at any one of positions 1 to 6 and / or any one of positions 8 to 11, and / or(ii) an amino acid substitution of Y at position 7 to any amino acid, in particular canonical amino acid, except L.

5. The antibody or antigen binding fragment thereof of any one of claims 1 to 4, wherein the variant of SEQ ID NO: 6 has(i) at most two, in particular one, amino acid substitutions at positions 1, 2, any one of positions 4 to 10 and / or 12, and / or(ii) an amino acid substitution of A at position 3 to any amino acid, in particular canonical amino acid, except L or I, or an amino acid substitution of N at position 11 to any amino acid, in particular canonical amino acid, except W.

6. An antibody or antigen binding fragment thereof binding to Hemoglobin Ale (HbAlc) comprising a heavy chain variable domain (VH) as shown in SEQ ID NO: 25 or variant thereof, and a light chain variable domain (VL) as shown in SEQ ID NO: 26 or variant thereof.

7. The antibody or antigen binding fragment thereof of any one of claims 1 to 6, wherein the antibody or antigen binding fragment thereof binds to an epitope comprising or consisting of the sequence as shown in SEQ ID NO: 33 or a variant thereof with one or two amino acid substitutions.

8. The antibody or antigen binding fragment thereof of any one of claims 1 to 7, wherein when the antibody or antigen binding fragment thereof is used in a turbidimetric inhibition immunoassay for determining the levels of HbAlc using a sample comprising a normal physiological concentration of HbAlc, the immunoassay has a coefficient of variation of 1.5 % or lower.

9. The antibody or antigen binding fragment thereof of any one of items 1 to 8, wherein when the antibody or antigen binding fragment thereof is used in a turbidimetric inhibition immunoassay for determining the levels of HbAlc using a sample comprising an abnormal physiological concentration of HbAlc, the immunoassay has a coefficient of variation of 1.5 % or lower.

10. A nucleic acid coding for at least a part of an antibody or antigen binding fragment thereof of any one of claims 1 to 9.

11. A composition comprising an antibody or antigen binding fragment thereof of any one of items 1 to 9 and an acceptable carrier.

12. Use of an antibody or antigen binding fragment thereof of any one of items 1 to 9, a composition of claim 11 for determining the amount or concentration of HbAlc in a sample.

13. Method of determining the ratio of HbAlc in a sample comprising the following steps:(a) adding to the sample an antibody or antigen binding fragment thereof of any one of claims 1 to 9,(b) measuring with a photometer the absorbance of the sample to determine the total amount of haemoglobin in the sample,(c) adding to the sample a polyhapten comprising at least one binding site for the antibody or antigen binding fragment thereof of any one of claims 1 to 9, and(d) measuring with a photometer the absorbance of the sample to determine the amount of HbAlc in the sample.

14. The method of determining the levels of HbAlc in a sample of claim 13, wherein the method has a coefficient of variation of 1.5 % or lower, when using a sample comprising a normal physiological concentration of HbAlc.

15. The method of determining the levels of HbAlc in a sample of claim 13 or 14, wherein the method has a coefficient of variation of 1.5 % or lower, when using a sample comprising an abnormal physiological concentration of HbAlc.

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