Improved antibody chip kit capable of quantitatively detecting multiple cell factors synchronously

A cytokine and antibody chip technology, applied in the field of biomedicine, can solve the problems of less specimen consumption, cumbersome operation, and low sensitivity, and achieve the effects of reducing background interference, increasing stability, and increasing detection sensitivity

Active Publication Date: 2014-10-22
RAYBIOTECH INC GUANGZHOU +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] Aiming at the deficiencies in the prior art, the object of the present invention is to provide an improved body chip kit for quantitatively detecting multiple cytokines at the same time. The kit uses fluorescent detection signals to quantitatively detect dozens of cytokines simultaneously, overcomi

Method used

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  • Improved antibody chip kit capable of quantitatively detecting multiple cell factors synchronously
  • Improved antibody chip kit capable of quantitatively detecting multiple cell factors synchronously
  • Improved antibody chip kit capable of quantitatively detecting multiple cell factors synchronously

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0031] Example 1: Screening of the best antibody chip carrier.

[0032] Conventional antibody chips mostly use nitrocellulose membrane as the carrier. Because the nitrocellulose membrane has a multi-layer structure, it is difficult to clean the chip, so that the results of the chip fluctuate greatly. At the same time, because nitrocellulose membrane chips are not easy to operate on a large scale, the use of large-scale clinical samples is not widespread. Different manufacturers use different methods to fix the nitrocellulose membrane on the surface of the glass slide. Among them is Whatman's SS glass slide, which fixes 16 cells containing nitrocellulose membrane on a glass slide. In addition, Gentel has used processing The nitrocellulose material is coated on the surface of the glass slide to produce the PATH glass slide. In addition, in order to coat the surface of the glass slide with antibodies, we screened the carriers activated by different methods. Spot Cy3 and Cy5 labele...

Example Embodiment

[0034] Example 2: Preparation of an antibody chip kit for simultaneous quantitative detection of multiple cytokines.

[0035] In order to detect the presence of the corresponding cytokines in the sample, prepare slides fixed with specific antibodies against the following proteins: granulocyte-macrophage colony stimulating factor (GM-CSF), interferon gamma (IFNgamma), interleukin-2 (IL-2), Interleukin 4 (IL-4), Interleukin 5 (IL-5), Interleukin 6 (IL-6), Interleukin 8 (IL-8), Interleukin 10 (IL-10), Interleukin 13 (IL -13), tumor necrosis factor alpha (TNFalpha).

[0036] 1. Preparation of antibody:

[0037] Using specific antibodies against the proteins listed in Table 1, the source, concentration, and names of the proteins against which the antibodies are directed are specified in Table 1:

[0038] Table 1 The name of the antigen protein targeted by the specific antibody, the source of the antibody, and the concentration information

[0039]

[0040] 2. Preparation and storage of ant...

Example Embodiment

[0049] Example 3: Experiment for quantitatively detecting cytokines using the kit of the present invention.

[0050] 1. Complete drying of the slide chip

[0051] Take the slide chip out of the box, after equilibrating at room temperature for 20-30 minutes, open the packaging bag, remove the sealing strip, and then place the chip in a vacuum desiccator or dry at room temperature for 1-2 hours.

[0052] 2. Press image 3 Perform a serial dilution of the cytokine standard gradient as shown in

[0053] 2.1. Add 500μl of sample diluent to the small tube of cytokine standard mixture, and re-dissolve the standard. Before opening the vial, centrifuge it quickly, gently tap up and down to dissolve the powder, and mark this vial as Std1.

[0054] 2.2. Mark 6 clean centrifuge tubes as Std2, Std3 to Std7, and add 200μl of sample diluent to each tube.

[0055] 2.3. Extract 100μl of Std1 and add it to Std2 and mix gently, then extract 100μl from Std2 and add it to Std3, so as to gradually dilute to...

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Abstract

The invention discloses an improved antibody chip kit capable of quantitatively detecting multiple cell factors synchronously. The kit includes a standard tissue slide which is coated with activated amino groups and is used as a surface carrier. Various antigen-specificity antibodies are spotted on a surface of the slide to form a microarray through a non-contacting sample spotting instrument. The slide is divided into sixteen small zones, which are not interfered with each other, through a detachable 2*8-hole plastic frame, wherein each small zone contains an antibody microarray. Each captured antibody is repeated by four times in each antibody microarray. The slide and the frame are tightly clamped with each other through plastic clamping plates which are arranged at the two sides to form an antibody chip. Experimental reagents of a multiple sandwich ELISA reaction are completed on the surface of the slide. By means of the antibody chip kit, multiple cell factors can be quantitatively detected synchronously. A sensitivity of the kit is similar to that of single ELISA but a dynamic detection range of the cell factors is wider. Repeated data, which is as four times as that of ELISA, can be obtained from a single-sample experiment. The kit is high in sensitivity, is high in flux, is little in sample usage amount, is low in cost and is easy to popularize.

Description

technical field [0001] The invention belongs to the technical field of biomedicine and relates to an antibody chip kit for quantitatively detecting multiple cytokines simultaneously. technical background [0002] Cytokines are small molecular polypeptides (low molecular weight proteins) that are synthesized and secreted by the body's immune cells and non-immune cells and regulate a variety of cellular physiological functions. [0003] Cytokines have a wide range of biological activities, including promoting the proliferation and differentiation of target cells, enhancing anti-infection and cell killing effects, promoting or inhibiting the expression of other cytokines and membrane surface molecules, promoting inflammatory processes, and affecting cell metabolism. Cytokines related to immunity mainly include lymphokines produced by lymphocytes, monokines produced by monocytes and macrophages, interleukin (IL), interferon (IFN), colony stimulating factor (colony stimulating fa...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N21/64
CPCG01N21/6402G01N33/68
Inventor 黄若磐毛应清
Owner RAYBIOTECH INC GUANGZHOU
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