Homogeneous immunometric fluorescent compound set for quickly and quantificationally detecting heart fatty acid binding-proteins (FABP) and preparation method of homogeneous immunometric fluorescent compound set

A fatty acid binding and immunological reagent technology, applied in the field of medical testing, can solve the problems of non-excitation, low fluorescence value, low non-specific fluorescence, etc., and achieve the effects of low cost, good specificity and simple operation

Active Publication Date: 2015-04-29
SHENZHEN AIRUI BIO TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, Eu 3+ There is a large difference between the maximum emission wavelength of fluorescent substances and Alexa647, and the background fluorescence value without antigen-antibody reaction is ver...

Method used

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  • Homogeneous immunometric fluorescent compound set for quickly and quantificationally detecting heart fatty acid binding-proteins (FABP) and preparation method of homogeneous immunometric fluorescent compound set
  • Homogeneous immunometric fluorescent compound set for quickly and quantificationally detecting heart fatty acid binding-proteins (FABP) and preparation method of homogeneous immunometric fluorescent compound set
  • Homogeneous immunometric fluorescent compound set for quickly and quantificationally detecting heart fatty acid binding-proteins (FABP) and preparation method of homogeneous immunometric fluorescent compound set

Examples

Experimental program
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Effect test

Embodiment 1

[0042] 1. Preparation of anti-FABP for labeling:

[0043] Purified monoclonal antibody against cardiac fatty acid binding protein expressed by genetic engineering was selected. Eu 3+ The product code of anti-cardiac fatty acid binding protein monoclonal antibody for labeling is 10E1; the product code of anti-cardiac fatty acid binding protein monoclonal antibody for fluorescein labeling is 9F3 and 5B5.

[0044] 2. Preparation of rare earth element chelate labeled anti-FABP:

[0045] The mouse anti-human H-FABP monoclonal antibody 10E1 solution (3 mg / ml) was dialyzed twice at 4° C. with 3 L of 0.9% NaCl, 24 hr each time. Add water to adjust the concentration to 1.5mg / ml. Take 0.6ml of the antibody solution, add 1ml NaHCO 3 (0.2mol / L), and adjust the pH to 9.1 with 1mol / L NaOH. 20 μl of BHHCT methanol solution (30 μg / ml) was added dropwise to the antibody solution under stirring, and the stirring reaction was continued for 1 hr. After centrifugation (10000rpm, 10min) to re...

Embodiment 2

[0051] The preparation method of the present embodiment is basically the same as that of Example 1, the difference is:

[0052] In step 2, the preparation method of rare earth element chelate-labeled anti-FABP is: dialyze the mouse anti-human H-FABP solution (3 mg / ml) twice at 4° C. with 3 L of 0.9% NaCl, 24 hr each time. Add water to adjust the concentration to 1.5mg / ml. Take 0.6ml of the antibody solution, add 1ml NaHCO 3 (0.2mol / L), and adjust the pH to 9.1 with 1mol / LNaOH. 20 μl of BHHBCB methanol solution (30 μg / ml) was added dropwise to the antibody solution under stirring, and the stirring reaction was continued for 1 hr. After centrifugation (10000rpm, 10min) to remove insoluble matter, put on SephadexG-25 column, use 0.05mol / L NH 4 HCO 3 (pH8.0) to separate the labeled protein and free label. UV / visible spectrophotometer detects the A of each collection liquid 330 value, pool the solutions containing the labeled antibodies. Add final concentrations of 0.1% BSA ...

Embodiment 3

[0054] The preparation method of the present embodiment is basically the same as that of Example 1, the difference is:

[0055] In step 3, the anti-H-FABP monoclonal antibodies 9F3 and 5B5 were diluted to 1mg / ml with 0.1M sodium bicarbonate solution respectively, 5ml of the antibody solution was taken respectively, and 40mg of fluorescein DyLight-DY647 solution was added, and stirred evenly. Incubate at room temperature for 1.5 hours, mixing every 15 minutes. Finally, use a G25 gel column to separate and purify, collect the labeled fluorescein-labeled antibody, dilute it with 0.01M phosphate buffer containing 0.025% PEG, 2.5% BSA, 15% glycerol, and 0.03% surfactant. The bottle is sealed and stored at 4°C.

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Abstract

The invention provides a homogeneous immunometric fluorescent compound set for quickly and quantificationally detecting heart fatty acid binding-proteins (FABP) and a preparation method of the homogeneous immunometric fluorescent compound set. The homogeneous immunometric fluorescent compound set comprises a rare-earth element chelate marked anti-FABP monoclonal antibody, a near infrared fluorescent compound marked anti-FABP monoclonal antibody and FABP calibrators with series concentration. The homogeneous immunometric fluorescent compound set can be used for detecting the low-value FABP and the high-value FABP, particularly the low-value FABP, is low in cost, simple, quick and sensitive to operate and good in specificity, only needs to be matched with a special homogeneous fluoroimmunoassay detection instrument, and therefore, the homogeneous immunometric fluorescent compound set can be widely applied to medical examination places at different levels, particularly basic-level medical mechanisms including health clinics in towns and townships and has great significance on preventing heart and cerebral vessel events.

Description

technical field [0001] The invention belongs to the field of medical examination, and in particular relates to a homogeneous fluorescent immunological reagent for rapid and quantitative detection of cardiac fatty acid binding protein and a preparation method thereof. Background technique [0002] The morbidity and mortality of acute myocardial infarction (AMI) are both high. Modern therapeutics believes that it is very important to quickly diagnose and perform reperfusion therapy after the onset of AMI to reduce mortality and improve prognosis. The diagnostic criteria for AMI recommended by WHO are: clinical symptoms, abnormal electrocardiogram and biochemical indicators. However, about 1 / 3 of AMI patients have atypical early clinical symptoms, and about 50% of patients cannot have characteristic ST-segment changes in ECG. Therefore, a sensitive and specific myocardial index is particularly important for the diagnosis of this period. [0003] Heart fatty acid binding protei...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/533
CPCG01N33/533G01N33/68
Inventor 谢爱武
Owner SHENZHEN AIRUI BIO TECH
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