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Compositions and methods for detecting albumin

A technology of albumin and protein ester, which is applied in the field of medical diagnosis and can solve the problem of probes limiting the clinical application of quantitative albumin

Pending Publication Date: 2021-05-21
图纳蒙特生物创业有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the specificity of the probe limits its clinical application to quantify albumin in biological samples

Method used

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  • Compositions and methods for detecting albumin
  • Compositions and methods for detecting albumin
  • Compositions and methods for detecting albumin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0062] This example demonstrates albumin and non-albumin esterase activity in human serum.

[0063] Human sera were fractionated using superose-6 columns followed by 14:0 NPS-PC as a probe to identify esterase activity in the presence or absence of HSA inhibitors. Such as figure 1 As shown in , three major fractions of esterase activity were identified in the absence of HSA inhibitors. When the esterase activity of HSA was inhibited, the third peak disappeared, indicating that the last peak was the esterase activity of HSA. The first and second peaks are non-albumin esterase or lipase activity.

[0064] There are two advantages to using 14:0 NPS-PC as a probe: First, due to the unique phosphorylcholine (PC) structure that limits its specificity to only phospholipase A2 (PLA2), non-albumin esterase / lipid Enzyme / hydrolase activity is lower; second, it is highly hydrophobic and can displace bound fatty acid esters or lipids on albumin and thus reduce protein underestimation. ...

Embodiment 2

[0066] This example demonstrates a method for determining the amount of albumin in a biological sample.

[0067] Reactions were started by adding 110 μl of TBS reaction buffer (10 mM Tris-HCl, 150 mM NaCl, pH 7.4) containing 0.5M 14:0 NPS-PC and 5 mM EDTA to 20 μl of HSA solution in a well of a 96-well plate. Reactions were followed at a wavelength of 405 nm (absorbance) in a SPECTRAmax M5 plate reader.

[0068] Such as figure 2 As shown in , the absorption at a wavelength of 405 nm increases while 14:0 NPS-PC is hydrolyzed by HSA to release p-nitrophenol. The method does not require pre-incubation. Depending on the concentration of albumin, the reading time can be from 0.5 to 30 minutes. Typical read time is 3-5 minutes. The reaction time temperature may be ambient temperature.

Embodiment 3

[0070] This example demonstrates that 14:ONPS-PC is an effective substrate for measuring the pseudoesterase activity of albumin.

[0071] image 3 Michaelis-Menten kinetics of HSA pseudoesterase using 14:0 NPS-PC as a probe is shown. Such as image 3 As shown in , 14:0 NPS-PC has a relatively low Km in the pseudoesterase reaction of HSA, and thus the measurement of HSA concentration can be sensitive and rapid.

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Abstract

The present disclosure provides a method for determining the amount of albumin in a sample. In one embodiment, the method involves treating the sample with an esterase inhibitor that selectively inhibits non-albumin esterase activity; combining the sample with a selective substrate of albumin, which has a carboxylic ester bond, so that the carboxylic ester bond is cleaved to generate a hydrolysate; detecting the amount of the hydrolysate generated in a period of time; and determining the amount of the albumin in the sample based on the amount of the hydrolysate in the period of time.

Description

technical field [0001] The present disclosure relates generally to medical diagnosis. More specifically, the present disclosure relates to compositions and methods for the specific detection and quantification of albumin. Background technique [0002] Albumins are a family of non-glycosylated globular proteins commonly found in plasma. Serum albumin is the main protein in human plasma and performs various physiological functions, such as maintaining colloid osmotic pressure, transporting various biomolecules, and exerting antioxidant effects. [0003] Albumin assays are typically performed in clinical biochemistry laboratories. For example, urinary albumin has been widely used as an important biomarker in patients with renal impairment such as diabetes, hypertension, and acute glomerulonephritis after streptococcal infection. The normal range of albumin in adult serum is 34-54g / L, and low albumin may be associated with liver disease, nephrotic syndrome, burns, protein-los...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68C12Q1/44
CPCG01N33/6893G01N33/6827C12Q1/44G01N2333/765C12Q2334/10G01N31/22G01N21/31G01N33/68
Inventor 卓少秋
Owner 图纳蒙特生物创业有限责任公司