82 results about "Phycoerythrobilin" patented technology

The present present invention relates to a method of simultaneously extracting phycoerythrin and agar from seaweeds such as asparagus, etc in seaweeds biological chemical field. The concrete operation process is that cells of the asparagus frond are crushed. A phycobiliprotein crude extract is obtained from water soluble extracts after ammoniym sulfate precipitation. The crude extract is extracted by Phenyl-Sepharose expansion columns and phycoerythrin products are obtained. Q-Sepharose iron exchange columns are used to purify the phycoerythrin products to obtain purified phycoerythrin products. Then, the left asparagus residues after the phycoerythrin extract are used to extract the agar. The present invention can simultaneously obtain the phycoerythrin and the agar from the asparagus. And the purified phycoerythrin purity is larger than 2.954, and the yield is 0.097mg per gram of fresh algae. The yield of the obtained agar is 2.31percent (the algae is wetly weighted), which is equivalent to the yield of an agar direct extraction method. Therefore, the utilization ratio of the asparagus can be greatly improved by adopting the method.

The invention discloses a non-diagnostic method for detecting a suspension chip of PCR products. The chip mainly comprises coded microspheres, a biotinylated primer, a capture probe, and streptavidin-biotin-phycoerythrin, and the method comprises the following steps that: the capture probe is coupled with corresponding microsphere of each size respectively, a red laser excites classified fluorescent lights on aspherical substrate, and the types are determined according to different colors of the spherical substrate, wherein the biotinylated primer shows the a primer needs biotinylation labeling during PCR; the microspheres coupled with the probe can be specifically combined with a PCR product labeled by an amplified biotin; and the streptavidin-biotin-phycoerythrin is combined with a biotin on the PCR product captured on the microspheres, a green laser excites the phycoerythrin, and the number of the reported fluorescent molecules combined on the spherical substrate is measured and is used for indirectly determining the content of the PCR product combined on the spherical substrate.

The invention relates to a method for preparing high purity phycoerythrin, phycocyanin and allophycocyanin from the algae, belonging to the bioengineering extraction separation technology. The method is as follows: red algae or cyanobacteria is adopted as a raw material, and phosphate buffer is used as an extractant; after algae cells are broken, the raw material of the red algae or the cyanobacteria is salt dissolved, salted out and dialyzed in grade by ammonium sulfate, and crude extract of phycobiliprotein is obtained; the crude extract is in chromatography by a hydroxyapatite column for one time, is gradient eluted by the phosphate buffer, and is frozen and dried, and high purity phycoerythrin, phycocyanin and allophycocyanin are obtained. The purity of the crude extract of the phycobiliprotein prepared by the effective method in the earlier stage reaches the standard of pharmaceutical grade (A620/A 280 is more than 2.0), thereby simplifying the follow-up purifying procedures, so the high purity phycoerythrin, phycocyanin and allophycocyanin can be respectively obtained by only one time column chromatography. When the method is used to extract high purity phycobiliprotein from the fresh algae and the dry algae, the process is simple, the production cost is low, and the yield of the products is high. Thus, the method is suitable for mass production.

The invention discloses a method for detecting a system for quickly screening a fever pathogen, and mainly relates to detection of a tuberculosis antibody, a flu antibody, a bird flu antibody, a plague antibody and an SARS antibody in blood serum. The chip mainly comprises coding microspheres, a coating antigen, a detection antibody, a biotinylated antibody and a streptavidin-phycoerythrin. The method comprises the following steps that: the coating antigen and the coding microshperes are coupled; the detection antibody is respectively coupled with the corresponding microspheres in separate specificity; the red laser excites the classified fluorescence on the spherical substrate; the type is determined according to different colors of the spherical substrates, wherein the biotinylated antibody is combined with the detection antibody; and the streptavidin-phycoerythrin is combined with the biotin of the detection antibody captured by the microshperes; the green laser excites the phycoerythrin, the number of report fluorescence molecules combined on the spherical substrate is measured, and the number is used for indirectly determining the content of the detection antibody combined onthe spherical substrate, so that whether the pathogen infection exists is determined, and the aim of quickly screening the fever pathogen is achieved.

The invention relates to phycocyanin beta subunits fluorescent protein combined with phycoerythrobilin PEB, fusion protein formed by phycocyanin beta subunits fluorescent protein and streptavidin and a mutant thereof, the sequences include sequence 1, sequence 2, sequence 3 and sequence 4; furthermore, the invention discloses a method of the fluorescent protein fused with streptavidin for fluorescent immunological detection directly; phycocyanin beta subunit conserved cysteine residue can be not only combined with phycocyanobilin PCB by a thioether bond, but the fact that the phycocyanin beta subunit conserved cysteine residue can be combined with the phycoerythrobilin PEB by the thioether bond through the genetic engineering can be realized, so as to obtain the novel fluorescent phycocyanin, the spectroscopy of the protein is completely different from that of the phycocyanin beta subunits fluorescent protein combined with PCB, and the protein has high fluorescence efficiency; in addition, as the protein carries His-tag label, purification is not only convenient, but also the dissolubility of the protein can be improved; the protein is combined with the streptavidin to form the fusion protein for fluorescent immunological detection directly, thereby being beneficial to the application of the protein in all kinds of the field.

A novel suspension hybridization assay was used to determine nucleic acid copy number by flow cytometry. The assay was validated with low copy (lc) products ranging in length from 100 to 2304 bp conjugated to spectrally-distinct polystyrene microspheres. In the example provided herein, these conjugated microspheres were used as multiplex hybridization probes to detect homologous sequences in genomic DNA extracted from cytogenetic cell pellets and labeled with biotin-dUTP. Hybridization was detected with phycoerythrin-labeled streptavidin and analyzed by flow cytometry. Copy number differences were distinguishable by comparing the mean fluorescence intensities of test probes with a diploid reference probe in genomic DNA of patient samples and abnormal cell lines. The assay is capable of distinguishing a single allele and three alleles at a test locus from a biallelic reference sequence, regardless of chromosomal context. The assay is an improvement on previous methods which require prior amplification of locus-specific target DNA because, lc probes provide adequate specificity and sensitivity for accurate copy number determination of homologous targets. Because of its high sensitivity and accuracy, the assay is useful for determination of nucleic acid copy number for a variety of applications, including determination of genomic copy number in humans, animal models of disease and in solution, measurement of transcript levels, forensic DNA analysis, and quality control analysis in agriculture.

The developed reagent is three-color immunophenotyping reagent for measurement of CD4 positive lymphocytes in peripheral blood. The reagent contains 7-aminoactinomycin D (7-AAD) which intercalates into double stranded DNA and is easily excited at 488 nm. The fluorescence emission of 7-AAD has peak at 670 nm that can be detected with FL3 detector of flow cytometer. The 7-AAD, therefore, stains white blood cells and discriminates it from red blood cells. The reagent also contains fluorescein isothiocyanate (FITC) labeled CD4 monoclonal antibody and phycoerythrin (PE) labeled CD14 monoclonal antibody which are detected with FL1 and FL2 detectors of flow cytometer, respectively. The developed reagent can be used to measure number of CD4 positive lymphocytes in lymphocyte population and monitor monocyte contamination simultaneously. This reagent therefore provides more accuracy results of CD4 positive lymphocyte enumeration.

The invention provides fluorescent analogues of thyroid hormones T3 and T4. Also provided are assays for thyroid hormones utilizing the compounds of the invention. The assays of the invention are in both one-step and two-step formats and allow the determination of both T3 and T4 concentration in a sample.

The invention relates to a preparation method of a phycoerythrin ratio fluorescence sensor based on a magnetic molecular imprinted nucleus/shell polymer. A Fe3O4 magnetic nano-particleis used as the center, the surface of the Fe3O4 magnetic nano-particleis coupled with blue fluorescence emitting carbon quantum dots B-CDs, a SiO2 shell layer loaded with template molecule phycoerythrin generates onthe surfaces of the Fe3O4/B-CDs, and template molecule elution is conducted to prepare magnetic molecular imprinted nucleus/shell polymer Fe3O4/B-CDs/SiO2-MIPs. The polymer dispersion liquid fluorescence emission spectra under different phycoerythrin concentration are measured, the linear relation between the fluorescence emission peak intensity ratio I<phycoerythrin>/I<B-CDs> of the phycoerythrinto the B-CDs and the phycoerythrin molar concentration is fitted, and a phycoerythrin ratio fluorescence sensor is constructed. Compared with the prior art, the method is simple to operate and low incost, raw materials are easy to obtain, ratio signal interference resisting capacity is high, accuracy is high, flexibility and selectivity are high, and a novel ratio fluorescence sensor can be developed and used for efficient phycoerythrin detection.

The invention relates to a preparation method of fluorescence antibody for detecting a Newcastle disease virus and a solid-phase immunofluorescence detection assay kit. The preparation method comprises the following steps of: respectively deriving R-phycoerythrin (RPE) and an antibody resisting the Newcastle disease virus (NDV) by using a cross-linking agent SPDP (N-succinimidyl-3-(2-pyridyldithiol) propionate), cross-linking the derivatives in a proper molar ratio, and purifying through HPLC (High Performance Liquid Chromatography) to prepare an RPE marked NDV fluorescence antibody. The solid-phase immunofluorescence detection assay kit is formed from the fluorescence antibody, an NDV-resisting antibody, an agarose microsphere, diluted hydrochloric acid and a washing liquid. The assay kit comprises the following detection flows of: coating an activated microsphere with the antibody, washing, combining with a sample to be measured, washing, combining with the fluorescence antibody, fully washing, exciting by blue and green light in a fluorescence microscope, observing and judging the result. The prepared fluorescence antibody has the advantages of high yield, high purity, bright orange fluorescence and good stability; and the assay kit has signal enrichment action by using a spherical carrier and can increase detection sensitivity. The invention is applicable to the rapid detection of the Newcastle disease virus.

The present invention belongs to the field of fluorescent proteins in biotechnology, and particularly relates to a preparation method of a high fluorescence intensity recombinant phycobiliprotein concatermer. According to the preparation method, a streptavidin gene is linked to an allophycocyanin alpha subunit gene through a linker sequence; on the basis, one or a plurality of allophycocyanin alpha subunit genes are connected in series through linker sequences to form a fusion gene; and the fusion gene, a phycobiliprotein lyase gent and a phycoerythrobilin biosynthetic enzyme gene co-express in Escherichia coli to obtain the recombinant allophycocyanin concatermer with characteristics of biotin binding ability and high fluorescence intensity. According to the present invention, tn the immunofluorescence assay, the recombinant phycobiliprotein concatermer can achieve the strong fluorescent signal compared to the recombinant phycobiliprotein monomer; and the prepared recombinant phycobiliprotein concatermer can be adopted as the fluorescent marker for immunofluorescence detection in the field of biology and biomedicine.

A Spirulina maxima extract of the present invention and Allophycocyanin (APC), R-phycoerythrin (R-PE), and C-phycocyanin (C-PC), which are components of the Spirulina maxima extract, show an effect of inhibiting cell death and A2E (pyridinium bis-retinoid), oxidation due to blue light, and therefore can be usefully applied as an active ingredient in a composition for preventing and treating retinal disease.

The invention discloses a multi-fluorescent immunoassay method for rapidly distinguishing 6 types of poultry respiratory pathogens. The multi-fluorescent immunoassay method is simple to operate; a target amplified fragment is obtained through a PCR (Polymerase Chain Reaction); then an amplified product, fluorescence coded microspheres and streptavidin-phycoerythrin are hybridized; an MFI (Mean Fluorescence Intensity) value is read through a detector to distinguish viruses of different types. According to the method disclosed by the invention, avian influenza viruses, chicken infectious bronchitis viruses, chicken Newcastle disease viruses, chicken infectious laryngotracheitis viruses, mycoplasma gallisepticum and mycoplasma synoviae can be accurately detected at the same time; the multi-fluorescent immunoassay method has high specificity, high sensitivity and good repeatability. Compared with a traditional detection method, the method disclosed by the invention realizes simultaneous detection of a plurality of types of different target molecules in the same sample; the use amount of the sample is less; the method is simple and rapid to operate and the detection cost can be greatly reduced.

The invention discloses a multiple fluorescence immunoassay method for detecting mycoplasma gallisepticum and mycoplasma synoviae. The multiple fluorescence immunoassay method has the advantages that the operation is simple; a target amplified fragment is obtained through PCR (polymerase chain reaction), then the amplified fragment, fluorescent encoding microspheres, and streptavidin-phycoerythrin are hybridized, and an MFI (mean fluorescence intensity) value is read through a detection instrument, so as to distinguish different types of pathogens; the mycoplasma gallisepticum and mycoplasma synoviae can be simultaneously detected and identified; the specificity is strong, the sensitivity is high, the repeatability is good, and the like; multiple types of molecules with different purposes in the same sample can be simultaneously detected; the flexibility is good, and the type of to-be-detected pathogens can be increased or decreased on the basis according to requirements.

The invention relates to a preparation method of a phycoerythrin labeled polystyrene microsphere fluorescent probe. The method includes: conducting swelling treatment on seed microspheres by two-stepswelling to obtain surface carboxylated polystyrene microspheres, and coupling the carboxyl on the surfaces of the microspheres with phycoerythrin to obtain the phycoerythrin labeled polystyrene microsphere fluorescent probe. The polystyrene microsphere fluorescent probe prepared by the method provided by the invention has the advantages of strong stability, high sensitivity, difficult quenching,high safety factor, and no toxic or side effect.

The invention discloses plural gel containing phycoerythrin, a preparation method and application. The plural gel is phycoerythrin/collagen/carboxylic carbon nano tube/polyacrylamide plural gel. According to the plural gel disclosed by the invention, the phycoerythrin is prepared into a form of the phycoerythrin/collagen/carboxylic carbon nano tube/polyacrylamide plural gel through an appropriate preparation method, the phycoerythrin can be applied to sensitization batteries, and photoelectric converting efficiency is improved.

The invention discloses a primer group, a kit and a method for detecting six mouse viruses by multiple immunity fluorescence. The primer group is designed by the aid of multiple RT-PCR (reverse transcription-polymerase chain reaction) technologies and Luminex liquid chip technologies and can simultaneously detect six high-infection viruses such as TMEV, MHV, MNV, Reo-3, MVM and MPV of mice. By the aid of the primer group, target amplification products are acquired through specific PCR, the amplification products, fluorescence encoding microspheres and streptavidin-phycoerythrin are hybridized, and different types of viruses are distinguished when a mean fluorescence intensity value is read by a Luminex detector. The method has the advantages of rapidness, high efficiency, high specificity and sensitivity, good repeatability and the like and can be applied to quality monitoring, epidemiological investigation and early warning of test mice. The primer group is good in flexibility, and varieties detecting the viruses can be increased and decreased as required on the basis.

The invention relates to a preparation method of multiple fluorescently-labeled polystyrene microspheres, and the method comprises the steps of preparing surface-carboxylated polystyrene microspheresby two-step seed swelling polymerization, and combining the polystyrene microspheres with quantum dots to prepare quantum dot fluorescent microspheres, coupling carboxyl groups on the surface of the quantum dot fluorescent microspheres with phycoerythrin to obtain the multiple fluorescently-labeled polystyrene microspheres. The method has the advantages of being simple and low in raw material price, the prepared polystyrene microspheres have a stable structure, by use of double fluorescent labeling of the quantum dots and the phycoerythrin, the multiple fluorescently-labeled polystyrene microspheres have the advantages of remarkable performance characteristics, high safety factors in the preparation process, and no safety hazard to the human body.