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Method for measuring concentration of protein in solution by using glutaraldehyde to induce infrared fluorescence

A technology of protein concentration and infrared fluorescence, which is applied in the direction of fluorescence/phosphorescence, material excitation analysis, etc., can solve the problems of long operation time and high counting cost, and achieve the effect of reducing operation time, facilitating batch operation and reducing cost

Inactive Publication Date: 2010-09-29
NORTHWESTERN POLYTECHNICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In order to overcome the disadvantages of high counting cost and long operation time in the prior art, the present invention provides a method for measuring protein concentration, which is low in cost and easy to operate

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Embodiment 1: Carry out protein quantification using the method of the present invention

[0018] 1. Bovine serum albumin (BSA) with unknown concentration (experimental group) and known concentration (standard group) 1250ng / μL, 625ng / μL, 312.5ng / μL, 156.25ng / μL, 78.125ng / μL, 39.06ng / μL μL, 19.5ng / μL, 9.75ng / μL, 4.88ng / μL solution, 2μL of each sample was dropped on the nitrocellulose membrane;

[0019] 2. Dry the nitrocellulose membrane naturally;

[0020] 3. Put the whole nitrocellulose membrane into a normal saline solution with a glutaraldehyde concentration of 0.1%, heat the solution to 80°C and keep it warm for 30 minutes;

[0021] 4. Wash the nitrocellulose membrane with water for more than two times;

[0022] 5. Use an infrared laser scanning imaging system, select 680nm wavelength laser excitation, and detect the fluorescence intensity of the sample at a wavelength greater than 720nm of the excitation laser;

[0023] 6. Combine the known protein concentration ...

Embodiment 2

[0026] Embodiment 2: Carry out protein quantification using the method of the present invention

[0027] 1. Bovine serum albumin (BSA) with unknown concentration (experimental group) and known concentration (standard group) 625ng / μL, 312.5ng / μL, 156.25ng / μL, 78.13ng / μL, 39.06ng / μL, 19.5ng / μL, 9.75ng / μL, 4.88ng / μL, 2.44ng / μL solutions, 1μL of each sample was dropped on the nitrocellulose membrane;

[0028] 2. Dry the nitrocellulose membrane naturally;

[0029] 3. Put the whole nitrocellulose membrane into a phosphate buffered saline (PBS) solution with a glutaraldehyde concentration of 1%, heat the solution in a microwave oven, and react for 5 minutes;

[0030] 4. Wash the nitrocellulose membrane with phosphate buffer for more than two times;

[0031] 5. Use an infrared laser scanning imaging system, select 710nm wavelength laser excitation, and detect the fluorescence intensity of the sample at a wavelength greater than 780nm of the excitation laser;

[0032] 6. Combine th...

Embodiment 3

[0035] Embodiment 3: Carry out protein quantification using the method of the present invention

[0036] 1. Bovine serum albumin (BSA) with unknown concentration (experimental group) and known concentration (standard group) 2500ng / μL, 1250ng / μL, 625ng / μL, 312.5ng / μL, 156.25ng / μL, 78.125ng / μL , 39.06ng / μL, 19.5ng / μL, 9.75ng / μL, solutions, 0.5μL of each sample was dropped on the nitrocellulose membrane;

[0037] 2. Dry the nitrocellulose membrane naturally;

[0038] 3. Put the whole nitrocellulose membrane into an aqueous solution with a concentration of 10% glutaraldehyde, heat the solution to 40°C, and keep it warm for 60 minutes;

[0039] 4. Wash the nitrocellulose membrane with saline for more than two times;

[0040] 5. Use an infrared laser scanning imaging system, select 690nm wavelength laser excitation, and detect the fluorescence intensity of the sample at a wavelength greater than 740nm of the excitation laser;

[0041] 6. Combine the known protein concentration an...

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PUM

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Abstract

The invention discloses a method for measuring concentration of protein in solution by using glutaraldehyde to induce infrared fluorescence. The method comprises the following steps: dropwise adding protein solution in an experimental group to be tested and a plurality of protein solution with known concentration in a criterion group on nitrocellulose membrane; after drying, placing the obtained mixture into glutaraldehyde solution, and then washing after heating; and detecting fluorescence intensity of samples, and calculating the protein concentration in the experimental group according to a standard curve drawn by the known protein concentration and the fluorescence intensity and the fluorescence intensity of the protein samples in the experimental group. The method has lowered cost, improved speed and reduced operating time, thereby facilitating bulk operation.

Description

technical field [0001] The invention relates to a measuring method, especially a method for determining protein concentration. Background technique [0002] Protein concentration determination is a frequently used technique in life science research. There are currently many methods for protein concentration determination, but each has limitations: the classic UV spectrophotometric method has limited sensitivity; conventional BCA, Lowry methods, etc. involve relatively complicated reagent preparation; some methods, such as the publication number CN168774 The operation steps of the Chinese patent are complicated, and it is easy to introduce human error, resulting in a decrease in the accuracy of the measurement results; some methods, such as the European patent G01N33 / 68A12, the US patent US2010047913, etc., combine liquid chromatography and mass spectrometry, although it improves the detection Sensitivity and accuracy, but due to the expensive chromatography and mass spectro...

Claims

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Application Information

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IPC IPC(8): G01N21/64
Inventor 商澎李京宝何刚王亮骞爱荣
Owner NORTHWESTERN POLYTECHNICAL UNIV