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Protein content determination method for nerve growth factor and application thereof

A technology of nerve growth factor and determination method, which is applied in the field of double-antibody sandwich enzyme-linked immunoassay of mouse nerve growth factor, and can solve the problems of poor accuracy

Inactive Publication Date: 2006-06-21
ZHEJIANG YONGNING PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Although there have been some research results on the determination of nerve growth factor, the accuracy of its determination is poor.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] 1. After diluting the coated antibody with antibody diluent, prepare 5 μg / ml coated antibody with the coated solution, and coat it on a polystyrene microwell plate as an immobilized antibody, and make 3 wells For parallel experiments, add 100 μl to each well and seal overnight at 4°C;

[0047] 2. Discard the liquid in the well, wash with washing solution 200-300μl / well for 3-5 times, and remove the residual liquid in the well;

[0048] 3. Add 200 μl of blocking solution to each well and incubate at room temperature for 60 minutes;

[0049] 4. Repeat step 2, wash 5 times with 300 μl / well, and remove the residual liquid in the well;

[0050] 5. Add 100μl standard substance with a concentration of 12.5ng / ml to each well, and incubate at room temperature for 60-120min;

[0051] 6. Repeat step 4;

[0052] 7. Add 100 μl solution antibody (1 μg / ml) to each well and incubate at room temperature for 60-120 minutes;

[0053] 8. Repeat step 4;

[0054] 9. Add 100 μl of enzyme...

Embodiment 2-5

[0059] Repeat the operation steps of Example 1 to carry out Examples 2-5, except that the concentrations of the standards are respectively 0 ng / ml, 25 ng / ml, 50 ng / ml, and 100 ng / ml. Calculate the average value according to the data measured by the standard substance at each concentration, and draw a standard curve according to the average value obtained.

Embodiment 6-8

[0061] Repeat the operation steps of Example 1 to carry out Examples 6-8, except that the standard products therein are replaced by the tested samples 1-3 respectively. Using the obtained standard curve, the determination result of the nerve growth factor in the sample can be obtained according to the measured result.

[0062] The sample content can be calculated by the formula y (sample concentration) = a (intercept) + b (slope) × (OD 450 value) to calculate. The actual sample concentration needs to be multiplied by the sample dilution factor. The content determination results in samples 1-3 are shown in Table 1 below.

[0063] Numbering

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PUM

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Abstract

The invention discloses a double-antibody sandwich enzyme immune gauging method of mouse nerve growth factor, which detects whether the NGF exists and the content qualitatively or quantatively.

Description

technical field [0001] The invention relates to a method for measuring the content of nerve growth factor protein, in particular to a double-antibody sandwich enzyme-linked immunoassay method for mouse nerve growth factor. Background technique [0002] Nerve growth factor (NGF) is one of the most important biologically active molecules in the nervous system, and it is also the earliest discovered and most typical neurotrophic factor, which affects the survival and differentiation of certain neurons in the peripheral and central nervous systems. Usually, NGF is synthesized and secreted by the target tissue cells dominated by its effector neurons. It binds to the corresponding receptors on the nerve endings, enters the axon, and then transports to the cell body through the retrograde axoplasm. In this process, a series of biological effects are caused, such as inducing neurite growth, promoting the synthesis of certain proteins and enzymes, and so on. [0003] Although there ...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/52G01N33/535
Inventor 陆仙芸林德君
Owner ZHEJIANG YONGNING PHARMA
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