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An antibody chip kit for detecting cytokines related to bone metabolism

A technology of cytokines and antibody chips, applied in the biological field, can solve the problems of not being able to monitor changes in the metabolic level of bone cells, and achieve the effect of overcoming low throughput and single detection indicators

Active Publication Date: 2019-10-29
RAYBIOTECH INC GUANGZHOU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Bone density detection is a predictive indicator of fractures. Measuring bone density at a specific site can predict the risk of local fractures, but it cannot monitor changes in bone cell metabolism.

Method used

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  • An antibody chip kit for detecting cytokines related to bone metabolism
  • An antibody chip kit for detecting cytokines related to bone metabolism
  • An antibody chip kit for detecting cytokines related to bone metabolism

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1: Screening of antibody chip carriers.

[0022] Conventional antibody chips mostly use nitrocellulose membrane as a carrier. Since the nitrocellulose membrane is a multi-layer structure, it is difficult to wash the chip, resulting in high background of the chip and large fluctuations in the results. At the same time, the nitrocellulose membrane is fragile, not easy to operate on a large scale, and it is not widely used for large-scale clinical samples. Traditional glass slides are cheap and have low background, which has brought breakthroughs in the widespread use of diagnostic and research fields. We screened the glass slides on the market that were not treated with the active method, and the spotting effect of the glass slides, whether it was aldehydes or aminations, was unstable. After a lot of screening, it is found that the temperature and the active reagent components on the surface of the slide are the key to the effect of spotting on the slide.

[002...

Embodiment 2

[0034] Example 2: Preparation of the antibody chip kit described in the present invention.

[0035] The antibody chip kit of the present invention comprises the following components:

[0036] (1) Quantitative antibody chip slide combined with 31 kinds of specific antibodies.

[0037] (2) Sample diluent: a common sample diluent. 2 vials of 15ml 5X concentrated diluent D for diluting samples, 1 vial of 15ml 5X concentrated diluent B for diluting antibody and HRP-streptavidin. 1X dilution B is 15mM, PBS buffer solution of pH7.4, solute and its mass concentration or molar concentration or volume concentration in said dilution B are as follows: 0.5% casein, 2-4% sucrose, 150mM NaCl . The 1X dilution D is 15mM, pH6.5 PBS buffer solution, the solute and its mass concentration or molar concentration or volume concentration in the sample dilution solution are as follows: 2-4% sucrose, 150mM NaCl.

[0038](3) Washing liquid: it is a general washing liquid. 20X Concentrated Wash Sol...

Embodiment 3

[0047] Example 3: The experiment of quantitative detection of cytokines related to bone metabolism by using the kit of the present invention.

[0048] 1. Complete drying of the slide chip

[0049] Take the slide chip out of the box, and after equilibrating at room temperature for 20-30 minutes, open the packaging bag, peel off the sealing strip, and then place the chip in a vacuum desiccator or dry at room temperature for 1-2 hours.

[0050] 2. Perform gradient dilution on the bone metabolism-related cytokine protein gradient shown in Table 1

[0051] 2.1 Add 500 μl of sample diluent to the vial of cytokine standard mixture and redissolve the standard. Before opening the vial, give it a quick centrifuge and gently pipet up and down to dissolve the powder. Label this vial as tube 1.

[0052] 2.2 Mark six clean centrifuge tubes as tube 2, tube 3 to tube 7, and add 200 μl of sample diluent to each small tube.

[0053] 2.3 Take 100 μl of tube 1 and add it to tube 2 and mix gent...

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Abstract

The invention relates to an antibody chip kit for detecting bone metabolism-related cytokines. The antibody chip kit comprises a solid-phase vector, a sample diluting solution, a cleaning solution, a mixture of standard freeze-dried protein substances, a detection antibody mixture, a Cy3 dye substrate bonded with streptavidin and a sealing film, wherein the solid-phase vector is a standard tissue slide coated with a specific antibody, and the specific antibody is targeted to 31 bone metabolism-related cytokines. According to the invention, the standard tissue slide is used as a surface vector, multiplex sandwich ELISA reaction can be completed on the surface of the glide, and high-flux sample operation can be carried out at the same time; and the disadvantages of a single detection index, consumption of time, labor and materials and the like of conventional ELISA in receptor detection are overcome.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to an antibody chip kit, in particular to an antibody chip kit for cytokines related to bone metabolism. Background technique [0002] Bone is a tissue constantly undergoing a metabolic activity of remodeling, involving two opposing processes, bone formation (osteoblasts) and bone resorption (osteoclasts). Under normal circumstances, bone resorption and bone formation are closely related to different hormones (such as PTH, vitamin D, steroids, and calcitonin, etc.) and internal regulatory factors (such as cytokines and growth factors, etc.). The bone resorption process is regulated by RANKL and M-CSF and inhibited by OPG. Bone formation is induced by many biological factors, especially BMPs, FGF, PDGF, and TGFβ; it is also regulated by M-CSF, ALP, osteocalcin, osteopontin, and osteonectin. Imbalances in bone resorption and bone formation lead to bone metabolic diseases such as osteoporo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/543
CPCG01N33/543
Inventor 黄若磐毛应清
Owner RAYBIOTECH INC GUANGZHOU