Method for detecting activity of lectin of beans

A technology for lectins and beans, which is applied in the field of measuring the activity of bean lectins, can solve the problems of large error in results, difficult operation, high subjectivity, etc., and achieves high stability and accuracy, simple operation method, and high measurement results. accurate effect

Inactive Publication Date: 2009-03-11
CHINA AGRI UNIV
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Problems solved by technology

[0005] The existing methods for measuring lectin activity mainly have the following disadvantages: (1) the equipment used is expensive, the operation steps are cumbersome, and the operation is difficult; (3) only qualitative or semi-quantitative assays can be performed; (4) standard cell suspensions are prepared The methods are different, the reproducibility is poor, and many methods require

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  • Method for detecting activity of lectin of beans
  • Method for detecting activity of lectin of beans
  • Method for detecting activity of lectin of beans

Examples

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[0038] Example 1

[0039] Determination of Mung Bean Lectin Activity

[0040] (1) Preparation of mung bean lectin solution

[0041] Take 25g of mung bean, crush it and pass it through a 40-mesh sieve, soak in 100ml PBS buffer solution (0.05M, pH7.0, containing 0.15M NaCl), stir at 0℃ for 6h, then centrifuge at 4℃, 16000×g for 15min, Collect the supernatant, soak the resulting precipitate in PBS buffer solution (0.05M, pH7.0, containing 0.15M NaCl), and stir at 0°C for 3 hours, and then centrifuge at 4°C and 16000×g for 15 minutes to collect The two supernatants were combined to obtain mung bean agglutinin solution.

[0042] (2) Preparation of standard red blood cell solution

[0043]Take fresh rabbit blood added with 10% sodium citrate, centrifuge at 1277×g for 15 min, then suspend the collected red blood cell pellet in normal saline, and centrifuge again, repeat this 5 times until the supernatant is no obvious red; Add potassium phosphate saline buffer (pH 8.5, 5mM, 25ml per ml o...

Example Embodiment

[0052] Example 2

[0053] Comparison test of different lectin determination methods

[0054] Various measurement methods are as follows:

[0055] (1) Spectrophotometric method: the preparation of mung bean lectin solution and the preparation of standard red blood cell suspension are the same as those of the enzyme labeling method (see Example 1). The mung bean agglutinin solution prepared in Example 1 was serially two-fold diluted and then the following operations were carried out in sequence: 1ml each of the above solution was pipetted into 15 test tubes of 0.5×8cm, and then 1ml of the standard red blood cell solution prepared in Example 1 was added. Mix each test tube with a vortex mixer, then put it into the 0.5cm colorimetric cell with a light path and let it stand for 90-120 minutes, then carefully (minimize shaking) into the spectrophotometer to measure A 620nm . Then calculate according to the following formula

[0056] log x = log...

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Abstract

The invention discloses a method for determining the activity of bean agglutinin, which mainly comprises the steps that: bean agglutinin solution is prepared, standard erythrocyte solution is prepared, then the bean agglutinin solution is carried out series doubling dilution on the 96-hole elisa plate and the standard erythrocyte solution with the same volume is added to each hole, finally, the light absorption value under 620nm is determined by using an eliasa and the activity of the agglutinin is calculated according to the formula. The invention has accurate determination result, high stability and precision, and strong comparability, establishes a method for preparing standard red blood cells which can be stored and placed for a long time; the tested samples and standard erythrocyte suspension required by the method are few, and a plurality of samples can be determined simultaneously: in addition, the invention has simple operation method, can directly read the data by the eliasa, thus being applicable to determining the activity of the agglutinin of big samples.

Description

technical field [0001] The invention belongs to a method for measuring the activity of bean lectin. Background technique [0002] Lectin is a protein that can be synthesized and secreted by both animal cells and plant cells and can bind to sugar. It is named lectin because it can agglutinate red blood cells. Lectin has more than one sugar-binding site, so it can participate in cell recognition and adhesion, and connect different cells. [0003] Lectins are mainly used in the following aspects: (1) separation and purification of sugar-containing polymers and cells; (2) identification of microorganisms; (3) research on the distribution of glycoproteins and glycolipids on the cell surface; (4) identification of sugar chain structures (5) diagnosis of malignant tumor diseases; (6) identification of blood types; (7) directional delivery of drugs, etc. [0004] Currently, methods for measuring lectin activity include cell agglutination, Mancini precipitation, Kocourek affinity e...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N1/28
Inventor 沈群郇美丽
Owner CHINA AGRI UNIV
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