Method for detecting alkaline phosphatase in body fluid

A phosphatase and alkaline technology, applied to the detection of alkaline phosphatase in body fluids, based on the field of bioluminescence quantitative detection of alkaline phosphatase, can solve the problems of high price, high analysis cost, interference with enzyme and substrate reactions, etc., to achieve Improve sensitivity and accuracy, strong anti-interference ability, and high sensitivity

Active Publication Date: 2015-05-27
THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
View PDF4 Cites 7 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method is limited by many factors. For example, there are many substances in body fluids that interfere with the reaction between the enzyme and the substrate, which limits the use of the kit.
In addition, Roche P800 automatic biochemical analyzer and supporting biochemical kits and CL-7300 automatic bioc

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for detecting alkaline phosphatase in body fluid
  • Method for detecting alkaline phosphatase in body fluid
  • Method for detecting alkaline phosphatase in body fluid

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0048] Example 1

[0049] Using bioluminescence method to detect the content of alkaline phosphatase in serum

[0050] (1) Dilute normal human serum 10 times with pure water, and then add a series of different concentrations of alkaline phosphatase to the serum so that the final concentration is 7.5×10 -2 , 3.75×10 -2 , 7.5×10 -3 , 3.75×10 -3 , 7.5×10 -4 , 3.75×10 -4 , 7.5×10 -5 , 3.75×10 -5 , 7.5×10 -6 , 3.75×10 7 And 1.85×10 -7 .

[0051] (2) Take 1000μL of serum sample, add 100μL of adenosine triphosphate solution to it and resuspend it, react for 30min at 37℃; then add 50μL of bioluminescence reagent (luciferin and luciferase), and measure the bioluminescence value with a portable ATP detector .

[0052] Take the concentration of alkaline phosphatase as the abscissa and the change of bioluminescence intensity as the ordinate to construct a standard curve. The content of unknown alkaline phosphatase in the serum can be detected by the standard curve. figure 2 It can be seen that t...

Example Embodiment

[0053] Example 2

[0054] Linear relationship between adenosine triphosphate concentration and bioluminescence value

[0055] Dilute the adenosine triphosphate solution with Tris-HCl to concentrations of 0, 0.01, 0.1, 0.5, 1, 10, 20, 50, and 100μM. Take 50μL each of adenosine triphosphate with different concentrations, and then add bioluminescence containing luciferase and luciferin Reagent 50μL, mix well and quickly use a portable ATP detector to determine the bioluminescence value. A standard curve was constructed based on the relationship between luminescence value and ATP concentration. The linear relationship between adenosine triphosphate concentration and bioluminescence value is as image 3 Shown from image 3 It can be seen that as the concentration of ATP increases, the bioluminescence value becomes larger, which has a good linear relationship.

Example Embodiment

[0056] Example 3

[0057] The relationship between the content of alkaline phosphatase and the change of bioluminescence value

[0058] Dilute alkaline phosphatase with water to a concentration of 0,1.5×10 -5 , 3×10 -5 , 1.5×10 -4 , 3×10 -4 And 1.5×10 -3 U / mL, then take 100μL of alkaline phosphatase diluent, then add 50μL of 50μM ATP solution and mix well, react for 45min in a 37°C incubator, and finally add 50μL of bioluminescence reagent to mix well, pass the portable ATP detector Detect the bioluminescence value. From Figure 4 It can be seen that this method can detect 1.5×10 -5 U / mL alkaline phosphatase.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

PropertyMeasurementUnit
Particle sizeaaaaaaaaaa
Specific saturation magnetizationaaaaaaaaaa
Concentrationaaaaaaaaaa
Login to view more

Abstract

The invention relates to a method for detecting alkaline phosphatase in body fluid. The method comprises the following steps: triphosadenine (ATP) is used as a catalytic substrate of alkaline phosphatase (ALP), wherein triphosadenine is converted into adenosine in the presence of the catalysis of alkaline phosphatase; and luciferase can be used for catalyzing fluorescein to emit light under the condition that energy is provided by triphosadenine, and the light emitting intensity is in direct proportion to the content of ATP. When the quantity of ALP in a sample is more, more ATP is consumed so that the change amount of biological light emission is greater. Therefore, the content of alkaline phosphatase can be calculated by measuring the change amount of the light emitting intensity of the biological light emission. With the adoption of the method, 10-15M alkaline phosphatase can be detected; compared with the prior art, the method has the advantages that the sensitivity is high, the anti-jamming capability is strong, the operation is simple, and a detection instrument is easy to carry and is suitable for an analysis method of clinical diagnosis, and besides the advantages, the detection time can be greatly shortened and the detection cost is reduced.

Description

technical field [0001] The invention relates to the technical field of immune detection, in particular to a method for detecting alkaline phosphatase in body fluids, in particular to a method for quantitatively detecting alkaline phosphatase based on bioluminescence. Background technique [0002] Alkaline Phosphatase (Alkaline Phosphatase, ALP), also known as alkaline phosphatase, is a class of enzymes that can catalyze the hydrolysis of phosphate bonds under alkaline conditions. Common alkaline phosphatases include intestinal alkaline phosphatase, non-tissue-specific alkaline phosphatase, and placental alkaline phosphatase. Normally, the activity of alkaline phosphatase in stem cells and colon cancer cells is significantly increased, and it is used as a qualitative and quantitative indicator of the degree of differentiation of colon cancer cells. In addition, elevated serum alkaline phosphatase, termed hyperalkaline phosphataseemia, has been associated with malignant bile ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): G01N21/76
Inventor 蔣兴宇陈翊平吴景张晓青曹丰晶牛亚静查瑞涛
Owner THE NAT CENT FOR NANOSCI & TECH NCNST OF CHINA
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap