Universal rapid magnetic separation method and kit for pathogenic organism
A technology of pathogenic microorganisms and magnetic separation, applied in the direction of analysis materials, measuring devices, instruments, etc., can solve the problems of complex preparation process of immunomagnetic beads, insufficient amount of conjugated antibody, time-saving and labor-saving, etc., to achieve fast and accurate The effects of diagnosis, promotion of industrialization development, and reduction of work intensity
Image
Examples
Embodiment 1
[0034] Example 1 Preparation of universal bridged magnetic beads
[0035] 1. Prepare the following reagents
[0036] Reaction buffer: 0.05mol / L MES, pH5.0;
[0037] Washing solution: HEPES (hydroxyethylpiperazine ethanesulfonic acid) of 0.02mol / L, pH7.4;
[0038] Diluent: 0.01 mol / L, pH 8.0 PB (phosphate buffer, made of NaH 2 PO 4 ·2H 2 O, Na 2 HPO 4 ·12H 2 O preparation);
[0039] Blocking solution: 1mol / L NH4Cl;
[0040] EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride) activation solution: the concentration is 20mg / ml, prepared with the aforementioned reaction buffer;
[0041] NHS (N-hydroxysuccinimide) activation solution: the concentration is 20mg / ml, prepared with the aforementioned reaction buffer;
[0042] Preservation solution: 0.1% BSA (bovine serum albumin), 0.02% sodium azide, 0.1% Tween-20, prepared with 0.02mol / L HEPES;
[0043] Streptavidin solution: the concentration is 0.35mg / ml, prepared with 0.01mol / L, pH8.0 PB to dissolve streptav...
Embodiment 2
[0062] Example 2 Separation and Purification of Salmonella Paratyphi A by Specific Capture Magnetic Beads
[0063] 1. To prepare magnetic beads specific for Salmonella paratyphi A, follow the steps below:
[0064] a) Take the universal bridging magnetic beads prepared in Example 1 and dilute to 2 mg / ml with washing liquid; then take 1 ml of murine Salmonella paratyphi A monoclonal antibody diluted with washing liquid to 1 mg / ml, and mix with 1 ml of 2mg / ml universal bridging magnetic beads for mixing, and react at 37°C for 1 hour;
[0065] b) Step a) After the reaction is completed, use washing liquid to magnetically separate and wash 3 times, and then use washing liquid to dilute to 1 mg / ml to obtain specific capture magnetic beads.
[0066] 2. Capturing test of Salmonella paratyphi A
[0067] Follow the steps below:
[0068] a) Take newly amplified Salmonella paratyphi A (ATCC9150) and adjust to 10 with sterile washing solution 4 CFU / ml;
[0069] b) Take 1ml of the abov...
Embodiment 3
[0080] Embodiment 3 four kinds of pathogens are separated synchronously
[0081] 1. Preparation of specific capture magnetic beads
[0082] a) Get the universal bridging magnetic beads of Example 1 diluted to 2mg / ml by washing liquid, 1ml / tube, 4 tubes in total; , Shigella dysenteriae monoclonal antibody, Candida albicans monoclonal antibody, and Bacillus cereus monoclonal antibody were mixed, and reacted at 37°C for 1 hour;
[0083] b) Magnetic separation and washing with washing liquid for 3 times, and then diluting to 1 mg / ml to make specific capture magnetic beads.
[0084] 2. Synchronous separation test:
[0085] a) Mixed sample preparation: prepare a mixed bacterial solution with Salmonella paratyphi A, Shigella dysenteriae, Candida albicans, and Bacillus cereus, and dilute it with sterile washing liquid to make the A The concentrations of Salmonella paratyphi, Shigella dysenteriae, Candida albicans and Bacillus cereus were all 10 4 CFU / ml, divide the mixed bacterial...
PUM
| Property | Measurement | Unit |
|---|---|---|
| particle diameter | aaaaa | aaaaa |
| particle diameter | aaaaa | aaaaa |
Abstract
Description
Claims
Application Information
- IPC
- G01N33/569; G01N33/531; G01N33/543
- CPC
- G01N33/56911; G01N33/531; G01N33/54326
- Inventors
- 农高惠; 何林声