Nanofiber-containing lateral flow test platform for hba1c analysis

Abstract

The invention relates to a nanofiber coated lateral flow test system for the rapid, accurate and more sensitive determination of HbA1c levels in patients with Type 2 diabetes.

Description

[0001] NANOFIBER-CONTAINING LATERAL FLOW TEST PLATFORM FOR HbA1c ANALYSIS

[0002] Field of the Invention

[0003] The invention relates to a nanofiber coated lateral flow test system for the rapid, accurate and more sensitive determination of HbA1c levels in patients with Type 2 diabetes.

[0004] State Of The Art

[0005] HbA1c is a hemoglobin derivative that occurs when exposed to high blood sugar for a long time. In many recent methods, HbA1c measurement is recognized as the most reliable way to assess Type 2 diabetes. Therefore, rapid and continuous monitoring of HbA1c levels in patients is necessary to slow the progression of Type 2 diabetes.

[0006] In the state of the art, high-performance liquid chromatography for HbA1c detection (HPLC), affinity chromatography and capillary electrophoresis. However, these methods are not sufficiently feasible because they require large and expensive instruments, are time-consuming and costly.

[0007] Another reported topic for HbA1c detection is lateral flow kits (LFA) due to their porous nature, the ability to control surface properties on a large scale and easy biological modification according to the desired application. This is a frequently used system. However, due to sample interference effects, accurate and precise measurement of HbA1c is not fully realized. Because of all these shortcomings and problems, innovation is needed.

[0008] Aim of the Invention

[0009] The present invention relates to a lateral flow test system suitable for use in the determination of HbA1c levels in patients with type 2 diabetes, which meets the above-mentioned requirements, eliminates all disadvantages and brings some additional advantages. The primary aim of the invention is to develop an easy to manufacture, low cost, fast response time and more sensitive lateral flow test system for use in the meaningful measurement of HbA1c protein, which is important in the follow-up of type 2 diabetes.

[0010] Another aim of the invention is to increase measurement accuracy and sensitivity by using an electrospun membrane.

[0011] In order to fulfill the aforementioned objects, the invention is a lateral flow test system suitable for the determination of HbA1c levels in patients with Type 2 diabetes and includes the use of polycaprolactone (PCL) and polyethylene glycol (PEG) electrospun nanofibers membranes in this system.

[0012] The structural and characteristic features and all the advantages of the invention will be more clearly understood by the detailed description below, and therefore the evaluation should be made in consideration of this detailed description.

[0013] Figures Clarifying the Invention

[0014] Figure 1 -Schematic representation of the lateral flow system for the measurement of Hb1Ac

[0015] Figure 2- Images of the lateral flow system of the developed system

[0016] Figure 3- Analysis for Hb1Ac sample measurement using biological serum sample

[0017] Elements Helping to Understand Figures

[0018] 1 Serum sample

[0019] 2 Flow direction 3 Sample pad

[0020] 4 Conjugation pad

[0021] 5 Nitrocellulose membrane

[0022] 6 PCL / PEG nanofiber membrane

[0023] 7 Test line

[0024] 8 Control line

[0025] 9 Absorbent pad

[0026] Detailed Description of the Invention

[0027] In this detailed explanation, the invention is intended only for a better understanding of the subject matter and in no way in a way that does not create a limiting effect.

[0028] The subject of the invention, a lateral flow test system suitable for use in determining HbA1c levels in patients with type 2 diabetes, contains polycaprolactone (PCL) and polyethylene glycol (PEG) electrospun nanofiber membranes (6) in its most basic form. In a preferred embodiment of the invention, the developed system comprises 70-90% PCL and 10-30% PEG. In a preferred embodiment of the invention, PCL may be replaced by polylactic acid (PLA) and PEG may be replaced by polyethylene oxide (PEG), polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP).

[0029] In the inventive system for the measurement of HbA1c, polycaprolactone / polyethylene glycol (PCL / PEG) nanofiber membranes are used to reduce the effect of interfering species, especially in blood and serum (1 ), and to increase the selectivity and sensitivity of the system. Nanofibers are polymeric materials widely used in many biomedical applications due to their high surface area-to-volume ratio, high porosity and interconnected network structure and easy preparation. Electrospun nanofibers are used in LFA to regulate sample mobility due to their capillary structure, absorb interfering species and improve functional binding.

[0030] With the inventive system; a system has been created to eliminate the interference effects, especially lipid-induced interference effects that may come from blood and serum (1 ) samples by knitting a PCL I PEG membrane in the range of 0.5-1 cm in front of the blood conjugation pad. With this system, interference effects from blood were reduced and 5-10 times more sensitive analyzes were performed than the existing system (Figure 1-3).

[0031] PCL and PEG in the developed system are used in the preparation of electrospun nanofibers. The molecular weight of PCL is preferably between 40,000-80,000 g / mol. PCL reduces the interference effect by providing hydrophobic interaction. The molecular weight of PEG is preferably between 1 ,000-3,000 g / mol. PEG reduces the interference effect by providing hydrophilic interaction.

[0032] In an exemplary embodiment of the invention; Au NPs were synthesized by applying the most widely used citrate reduction method in the literature. The Au NP allows the test and control lines (8) to be seen clearly. Its size should be at most 40 nm. The synthesized Au NPs are conjugated with monoclonal anti-Hb antibody (detection antibody). Monoclonal anti-Hb antibody will be used as a detection antibody and must be specific to the antigen. AuNP / monoclonal anti- Hb antibody conjugates are impregnated into the conjugation pad. The conjugation pad (4) should be blocked and suitable for sample flow. Monoclonal anti-HbA1c antibody and goat anti-mouse IgG antibody are immobilized on nitrocellulose membrane. The monoclonal anti-HbA1c antibody is used for the test line and must be antigen specific. Goat anti-mouse IgG antibody will be used for the control line and should be antigen specific. PCL / PEG nanofibers are coated on the nitrocellulose membrane on the bottom of the conjugation pad (4) of the prepared lateral flow platform. The lateral flow platform was prepared to consist of a sample application pad, conjugation pad (4), nitrocellulose membrane and an absorbent pad (9). The nitrocellulose membrane should be blocked and suitable for sample flow. Sample pad (3); blocked and suitable for sample flow. Adsorbent pad; blocked and suitable for sample flow.

[0033] The invention starts by dropping the liquid containing the biological sample onto the sample pet (3) and

[0034] The test is completed in about 10 minutes. The following points should be noted:

[0035] - The sample volume applied to the sample pad (3) should not be too large.

[0036] - The Au NPs should be at most 40 nm in length.

[0037] - The thickness of the nanofiber coating needs to be considered.

[0038] Figure 1 is a schematic representation of the lateral flow system for Hb1Ac measurement developed in this study. Here, 1 indicates serum sample, 2 indicates flow direction, 3 indicates sample pad, 4 indicates conjugation pad, 5 indicates nitrocellulose membrane, 6 indicates PCL / PEG nanofiber membrane, 7 indicates test line, 8 indicates control line, 9 indicates absorbent pad. Figure 2 shows images of the developed lateral flow system. A. Example lateral flow system and 1 shows the part where the nanofiber coating is made, B-E. shows the top and side view of the PCL / PEG nanofiber membrane coating. Analysis for Hb1Ac sample measurement using biological serum sample in the system developed in Figure 3, 1. Control lateral flow system 2. 3. 2 min uncoated lateral flow system with PCL / PEG membrane, 4. 3 min uncoated lateral flow system with PCL / PEG membrane 5. 4 min uncoated lateral flow system with PCL / PEG membrane 6. 5 min uncoated lateral flow system with PCL / PEG membrane.

Claims

CLAIMS1. Lateral flow assay system for use in the determination of HbA1c levels in patients with type 2 diabetes flow testing system, is characterized by including polycaprolactone (PCL) and polyethylene glycol (PEG) electrospun nanofiber membranes (6).

2. The width of the nanofiber coating is in the range of 0.5-1 cm and the coating thickness is be in the range of 1-10 minutes depending on the nanofiber knitting time.

3. A product according to claim 1 , characterized in that it is composed of 70-90% PCL, 10-30 contains an amount of PEG in between.

4. A product according to claim 1 , characterized in that instead of PCL, it contains polylactic acid (PLA) and instead of PEG, it contains polyethylene oxide (PEG), polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP).

Images