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An immunomagnetic bead chromatography test strip and rapid detection method for detecting Cronobacter

A technology of immunomagnetic bead chromatography and Cronobacter, applied in biochemical equipment and methods, coatings, analytical materials, etc., can solve the problems of complex operation, inability to record, low sensitivity, etc., to simplify the optimization process and save money. The effect of less time, materials and reagents

Active Publication Date: 2020-11-24
HEFEI UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, ELISA reagents require special laboratories and professionals to conduct inspections, and the operation is complicated and takes a long time; colloidal gold reagents are easy to operate, fast, and suitable for on-site detection, but the sensitivity is slightly low, and the results are directly observed with the naked eye, and errors will occur. nor record

Method used

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  • An immunomagnetic bead chromatography test strip and rapid detection method for detecting Cronobacter
  • An immunomagnetic bead chromatography test strip and rapid detection method for detecting Cronobacter
  • An immunomagnetic bead chromatography test strip and rapid detection method for detecting Cronobacter

Examples

Experimental program
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Effect test

Embodiment 1

[0065] Rapid detection method of artificially contaminated milk powder samples with Cronobacter

[0066] 1. Preparation of immunomagnetic bead chromatography test strips for detecting Cronobacter genes

[0067] See image 3 with Figure 4 The immunochromatographic test strip is composed of a bottom plate and a sample pad, a nitrocellulose membrane, and a water-absorbing pad that are sequentially connected on the bottom plate. On the nitrocellulose membrane, a linear invisible detection line T line printed with a capture probe is used. The straight line invisible control line C printed with serum albumin antibody solution.

[0068] (1) The detection T-line is made of 2.5 μL of 100 μM capture probe solution with lengths of 10 bp, 20 bp, 30 bp and 40 bp. The capture probe sequence is as follows:

[0069] 10bp: 5'-Biotin-GGCGGAGCCG-3'

[0070] 20bp: 5'-Biotin-GGCGGAGCCGAATAACTGGG-3'

[0071] 30bp: 5'-Biotin-ATGACGTGACCGGCCAGTATTTCATGCACC-3'

[0072] 40bp: 5'-Biotin-GGCGGAGCCGAATAACTGGGAATCG...

Embodiment 2

[0111] Rapid detection method of Cronobacter in actual infant milk powder samples

[0112] 1. Preparation of immunomagnetic bead chromatography test strips for detecting Cronobacter genes

[0113] See image 3 with Figure 4 , The immunomagnetic bead chromatography test strip is composed of a bottom plate and a sample pad, a nitrocellulose membrane, and a water-absorbing pad that are sequentially connected on the bottom plate. On the nitrocellulose membrane, a linear invisible detection T line printed with a capture probe is used. The straight and invisible control C line printed by the anti-bovine serum albumin antibody solution.

[0114] (1) The detection T line is mixed with 2.5 μL of 100 μM 20bp capture probe, 2.5 μL of 5 mg / mL streptavidin and 15 μL of 50 mM phosphate buffer (volume ratio 1:1:6). The reaction was shaken for 3 hours to prepare a capture probe solution, and the DNA sequence of the capture probe was: 5'-Biotin-GGCGGAGCCGAATAACTGGG-3'.

[0115] (2) The control line...

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Abstract

The invention relates to an immunomagnetic beads chromatography test paper strip for detecting cronobacter. A nitric acid fiber film of the test paper strip is provided with a detection line T and a control line C, the detection line T is prepared by a capture probe solution obtained by uniformly mixing a 100 [mu]M capture probe with a length being 20 bp, 5 mg / mL of streptavidin, and 50 mM of a phosphate buffer according to a volume ratio being 1:1:6; the control line C is prepared by a standard solution with concentration being 1-5 mg / mL which is obtained by uniformly mixing an anti-bovine serum albumin antibody solution and a phosphoric acid buffer. when the immunomagnetic beads chromatography test paper strip is used for detecting whether milk powder contains cronobacter or not, detected liquid is prepared by bacteria DNA extraction, polymerase chain reaction, and PCR product and immunomagnetic beads combination on detected milk powder; when the detected liquid is dropped on a sample pad of the immunomagnetic beads chromatography test paper strip, a reaction at the room temperature is carried out for 10 min to obtain the observation result. If the detection line T and the control line C display yellow lines, the detected liquid is positive; if the control line C displays yellow, the detected liquid is negative, and the detection reaction time of a practical sample is shortened to 8-11 h.

Description

Technical field [0001] The invention belongs to the technical field of Cronobacterium gene detection, and specifically relates to an immunomagnetic bead chromatography test strip for rapid detection of Cronobacterium gene and a preparation method thereof. Background technique [0002] Cronobacterium (also known as Enterobacter sakazakii) is a species of Enterobacteriaceae, which can cause severe neonatal meningitis, enterocolitis and bacteremia, and the mortality rate is as high as 50%. Cronobacterium has attracted the attention of relevant departments in many countries around the world. According to reports, foreign dairy giants have been recalled for this reason. After the US FDA in 2002 detected Cronobacterium in infant formulas produced by some international dairy giants in the country, another international dairy giant voluntarily recalled a batch of products produced in the United States that had detected very small amounts of Cronobacter in 2003. Special formula milk pow...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6804G01N33/569G01N33/561G01N33/558G01N33/543
CPCG01N33/54326G01N33/54346G01N33/559G01N33/561G01N33/56916G01N2333/265G01N2446/90
Inventor 叶应旺聂文芳陈伟滕军徐建国姚丽
Owner HEFEI UNIV OF TECH
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