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Specific candida albicans fluorescent staining solution as well as preparation method and application thereof

A fluorescent dyeing and specific technology, applied in fluorescence/phosphorescence, preparation of test samples, material analysis through optical means, etc., can solve the problems of inability to specifically identify Candida albicans, easy inactivation of biological enzymes, and difficulty in preservation, etc. question

Pending Publication Date: 2021-09-10
广州浩康生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Later, a luciferin kit coupled with recombinant chitinase appeared on the market to detect fungi. This method has improved specificity and stability, but biological enzymes are easy to inactivate and difficult to store, and chitinase detection method is widely used. Spectrum fungal detection method cannot specifically identify Candida albicans

Method used

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  • Specific candida albicans fluorescent staining solution as well as preparation method and application thereof
  • Specific candida albicans fluorescent staining solution as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] The preparation of embodiment 1 streptavidin-fluorescein isothiocyanate complex

[0047] Dissolve 50 μL of FITC (3 mg / mL) in Add carbodiimide hydrochloride and N-hydroxysulfosuccinimide to 50 μL of borate buffer solution with a pH value of 5.5, and react for 3 to 10 minutes to adjust the pH value of the solution to 8.0 to 8.5, and then Streptavidin was added according to the molar ratio of streptavidin to fluorescein isothiocyanate at a ratio of 20:1, after mixing, the reaction was continued for 2 hours, and 10 μL of phosphoric acid containing 5% bovine serum albumin (BSA) was added. Block the reaction with salt buffer solution (PBS) for 10 min to stop the reaction, centrifuge to remove the precipitate, wash with borate buffer solution with a pH value of 7.0 in an ultrafiltration centrifuge tube with a molecular weight cut-off of 100K to obtain streptavidin-isothiocyanate Acid fluorescein complex, and resuspend the streptavidin-fluorescein isothiocyanate complex in bor...

Embodiment 2

[0048] The preparation of embodiment 2 adhesion peptide stock solution

[0049] Wash the adhesion peptide powder with phosphate buffer containing dimethyl sulfoxide (including 8.0g / L sodium chloride, 0.2g / L potassium chloride, 1.44g / L disodium hydrogen phosphate, 0.24g / L diphosphate Potassium hydrogen, 1% (mass percentage) of dimethyl sulfoxide, and the pH value is 8.0) to prepare a stock solution, and the concentration of the adhesion peptide in the stock solution is 1 mg / mL.

Embodiment 3

[0051] Incubate the adhesive peptide stock solution with the streptavidin-fluorescein isothiocyanate complex at room temperature for 30 min, shake thoroughly, and store in the dark to obtain a complex solution, add 500 mL of purified water to the beaker, and The obtained complex solution, potassium dihydrogen phosphate, disodium hydrogen phosphate, glycerol and dimethyl sulfoxide were mixed evenly, and the volume was adjusted to 1 L to obtain the specific Candida albicans fluorescent staining solution. Contains: Adhesive peptide stock solution 0.05g, streptavidin-fluorescein complex 0.05g, potassium dihydrogen phosphate 10g, disodium hydrogen phosphate 20g, glycerol 7g, dimethyl sulfoxide 5g.

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Abstract

The invention discloses a specific candida albicans fluorescent staining solution as well as a preparation method and application thereof. Every 1 L of the specific candida albicans fluorescent staining solution contains 0.05 to 0.15 g of an adhesive peptide stock solution, 0.05 to 1 g of a streptavidin-fluorescein compound, 5 to 25 g of a penetrating agent, 30 to 225 g of a stabilizing agent and 0.5 to 25 g of an anti-quenching agent. According to the invention, water is used as a solvent, and all other components are uniformly dispersed in the water and can stably coexist in the same system, so that the staining solution is more convenient and faster in the production, sales and use processes; the specific candida albicans fluorescent staining solution provided by the invention can specifically detect candida albicans, and is high in stability; fluorescent groups in the staining solution are excited by blue light and emitted by green light, so that the reduction of ultraviolet damage is facilitated, and microscopic observation is facilitated; the preparation method provided by the invention has the advantages of simple steps and mild reaction conditions, and can be completed only by stirring at normal temperature.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a specific Candida albicans fluorescent staining liquid and its preparation method and application. Background technique [0002] In recent years, with the widespread application of broad-spectrum antibiotics, anti-tumor chemotherapy, immunosuppressants and other drugs, the incidence of opportunistic fungal infections, especially deep fungal infections, has shown a clear upward trend. Candida albicans is one of the most common opportunistic pathogenic commensal fungi. It is part of the healthy human microbiota and exists in the host's mouth, epidermis, gastrointestinal tract, and vagina. It can cause serious illness in immunocompromised patients. and frequent mucosal candidiasis, such as oral and vaginal candidiasis (VVC) or systemic candidemia. The condition develops rapidly after infection, and the fatality rate of Candida albicans infection in domestic general ho...

Claims

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Application Information

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IPC IPC(8): G01N1/30G01N21/64
CPCG01N1/30G01N21/6428G01N2001/302G01N2021/6439
Inventor 苏成康陶成龙
Owner 广州浩康生物科技有限公司
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