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Test paper for diagnosing premature rupture of fetal membrane and reagent kit

A technology of premature rupture of membranes and test strips, applied in the field of immune applications, can solve the problems of high requirements for operators, need to operate instruments, and low sensitivity, and achieve the effect of low temperature requirements, high sensitivity, and strong specificity

Inactive Publication Date: 2008-07-09
BIO SOURCE WUHAN TECH LT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Another patent name is disposable premature rupture of membranes pH test paper diagnostic stick (200620083734.1), the disadvantage of this invention is that the sensitivity is not high, and it is easy to cause false negatives
Another patent name is A MARKER FOR PROLONGED RUPTURE OF MEMBRANES (WO2007030890). The method for detecting amniotic fluid in this patent is the ELISA method. Compared with the colloidal gold immunochromatography method, the disadvantage of this method is that it takes a long time and requires operating instruments. higher

Method used

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  • Test paper for diagnosing premature rupture of fetal membrane and reagent kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Preparation of AMNI-Pr

[0022] Amniotic fluid samples of 25-975ml were obtained from women undergoing caesarean section and were aliquoted in 2ml portions into 5ml centrifuge tubes and filtered with glass wool before aliquoting.

[0023] Mix 20 tubes of amniotic fluid with each tube to prepare a sample pool, and then use the following method to purify: firstly use Affi-Gel blue glue (Aurum Affi-Gel Blue Mini Kit) to pre-treat to remove endogenous albumin, Affi-Gel blue glue is purchased The samples in the sample pool were pretreated according to the instructions of the purchased Affi-Gel blue glue from Bio-Rad, USA.

[0024] The samples treated with Affi-Gel blue gum were then treated with the immunochromatography method described in International Patent No. WO03 / 018634 to obtain amniotic fluid-specific proteins.

[0025] Finally, two kinds of molecular weight cut-offs are respectively 300kD and 100kD centrifugal ultrafiltration tubes (AmiconUltra-4). Finally, three ...

Embodiment 2

[0027] Preparation of anti-AMNI-Pr monoclonal antibody

[0028] Refer to the method in Xue Qingshan's "Principles and Techniques of In Vitro Culture" Science Press, 2001 edition: use the AF1, AF2 and AF3 prepared above to immunize BALB / c mice respectively, and BALB / c mice were purchased from Experimental Animals of Hubei Academy of Medical Sciences In the center, the immunization procedure is to emulsify 150 μg of AF1 or AF2 or AF3 protein solution with an equal volume of Freund’s complete adjuvant, which was purchased from Sigma, and injected into the abdominal cavity of mice. After 21 days, the booster immunization was replaced with Freund’s complete adjuvant. Freund's incomplete adjuvant was emulsified, and Freund's incomplete adjuvant was purchased from Sigma Company; finally, three days before the fusion, or more than 4 weeks after the first immunization time, the intraperitoneal booster immunization of BALB / c mice, The amount of antigen was doubled to 300 μg, no adjuvant...

Embodiment 3

[0030] 1. Preparation of Ascites

[0031] After pretreatment of BALB / c mice with sterilized liquid paraffin, ascites was produced by injecting hybridoma cells. The specific process is as follows:

[0032] Pretreatment of BALB / c mice: Treat mice with sterilized liquid paraffin, intraperitoneally inject 0.5 mL / mouse, and inject hybridoma cells ten days later.

[0033] Expansion culture of positive wells: add feeder cells to 24-well cell culture plate, 4 drops / well, transfer positive cells in 96-well plate to 24-well cell culture plate for expansion culture.

[0034] Injection: resuspend the cells in the 24-well plate until the cells cover the bottom of the well, centrifuge at 1200r / min for 5min, count the cells, dilute them with RPMI-1640 basal culture medium to 5×106 cells / ml, and inject 0.5mL of mice intraperitoneally / mouse, and ascites can be collected when the abdominal bulge of the mouse is observed in about 8 days.

[0035] 2. Pretreatment of Ascites

[0036] Centrifu...

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Abstract

The invention relates to a test strip and a kit for premature rupture of membrane. The test strip comprises, from top to bottom of the horizontal plane of the test strip, a water absorbent pad, a cellulose nitrate membrane, a gold standard pad and a sample pad, wherein a detection line and a quality control line are coated on the cellulose nitrate membrane; the detection line is a monoclonal antibody 11B6 and the quality control line is rabbit anti-mouse IgG antibody; a monoclonal antibody 14A3 marked by colloidal gold is coated on the gold standard pad; and the absorbent pad, the cellulose nitrate membrane coated with the detection line and the quality control line, the gold standard pad coated with the monoclonal antibody 14A3 marked by the colloidal gold and the sample pad are sequentially adhered on a water-nonabsorbent support thin sheet from top to bottom; and a sample injecting hole is provided on the sample pad. The test strip in the kit has the advantages of high specificity and high sensitivity, and can be used for clinic diagnosis of premature rupture of membrane. The kit has simple operation and can determine the result in ten minutes.

Description

technical field [0001] The invention belongs to the application field of immunization, and relates to related fields such as molecular biology, immunization, and medicine. Specifically, it relates to premature rupture of membranes and extravasation of amniotic fluid, and then detects specific markers in the overflowing amniotic fluid to diagnose premature rupture of membranes, a test strip and a kit. Background technique [0002] Premature rupture of membranes (PROM) is a common complication in obstetrics, which can easily lead to complications such as intrauterine infection and fetal distress, and increase perinatal mortality and morbidity, but there is no ideal method to accurately evaluate fetal Intrauterine safety. Rupture of membranes prior to delivery occurs in 6.6%-13.9% of all patients, and spontaneous abortion occurs in 70%-90% of these cases within the next 48 hours. Premature rupture of membranes not only directly or indirectly causes 4.5%-14% of premature birth...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/558G01N33/531
Inventor 王宇波
Owner BIO SOURCE WUHAN TECH LT
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