Real-time fluorescent quantitative PCR (polymerase chain reaction) kit for detecting candida glabrata

A Candida and fluorescence quantitative technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of time-consuming, expensive, complicated operation, etc., and achieve cost reduction, easy operation, and simple operation Effect

Inactive Publication Date: 2011-09-14
SOUTHERN MEDICAL UNIVERSITY
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

Ordinary PCR is rarely used in clinical testing because it is easy to cause laboratory contamination. Universal primer PCR has the advantage of being fast and sensitive, but the specificity of isolation is poor, especially for Candida glabrata, which is not sensitive to some antifungal drugs. identification of bacterial strains; nucleic acid probes require a large number of probes to increase specificity, which are expensive and time-consuming, and cannot be used in routine laboratory inspections; PCR-SSCP, PCR-RFLP, gene chip technology, etc. are relatively sensitive and specific Good, but the operation is more complicated, the equipment requirements are high, and it is mostly used for scientific research, and it is not suitable for clinical testing at present

Method used

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  • Real-time fluorescent quantitative PCR (polymerase chain reaction) kit for detecting candida glabrata
  • Real-time fluorescent quantitative PCR (polymerase chain reaction) kit for detecting candida glabrata
  • Real-time fluorescent quantitative PCR (polymerase chain reaction) kit for detecting candida glabrata

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Embodiment 1 Candida glabrata detection kit and its use

[0040] 1. Prepare a kit including the following components: 1 tube of primer-probe mixture (25 μl / tube), 1 tube of PCR reaction solution (250 μl / tube), 1 tube of PCR enzyme system (75 μl / tube), positive quality control product (200μl / tube) 1 tube, negative quality control product (200μl / tube) 1 tube, pure H 2 O (2000μl / tube) 1 tube. Positive quantitative standard 4 tubes (10 7 copies / ml, 10 6 copies / ml, 10 5 copies / ml, 10 4 copies / ml).

[0041] 2. Collection, preservation and transportation of specimens

[0042] 2.1 Specimen types: blood, sputum, alveolar lavage fluid, etc.

[0043] 2.2 Specimen collection, storage and transportation: Take 200ul of blood, sputum, alveolar lavage fluid, etc., put them in sterile centrifuge tubes, label them well, put them in a curling container and take them to the laboratory or at low temperature (-20℃~-70℃ ℃) frozen. Dry ice is used for long-distance transport of specim...

Embodiment 2

[0054] Example 2 Application of Candida glabrata detection kit to detect clinical samples

[0055] Select 3 cases identified as Candida glabrata negative through the culture method, 3 cases were identified as Candida glabrata positive specimens through the culture method, the nucleic acid extraction of the specimen, PCR amplification and result analysis steps were carried out with reference to Example 1, and at the same time Perform negative and positive quality control testing.

[0056] Test result: the amplification curve of the negative quality control product is not S-shaped (see figure 1 ), the amplification curve of the positive quality control product is an obvious S-shaped curve (see figure 2 ), both negative and positive quality control products meet the quality control requirements of the kit; the instrument draws a standard curve based on the positive quantitative standard (see image 3 ) has a good linear relationship, indicating that the amplification efficienc...

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Abstract

The invention discloses a real-time fluorescent quantitative PCR (polymerase chain reaction) kit for detecting candida glabrata, which comprises the following substances: PCR reaction solution, primer and probe mixed solution, a PCR reaction enzyme, pure H2O and a positive quantitative standard product. The real-time fluorescent quantitative PCR kit is used for detecting the nucleic acid amplification level of the candida glabrata and can reflect the state of the infection of the candida glabrata in a patient body, be used for monitoring, preventing and treating fungal infection, and be conductive to early diagnosis and treatment of diseases; and the real-time fluorescent quantitative PCR technology designs a primer and a probe against the nucleic acid specificity sequence of the candida glabrata, the specificity is strong, the sensitivity is high, and the technology further has the advantages of higher flux, simpleness in operation and relatively low cost, and is applicable to detecting various samples of most of patients. The amplification of the samples to be detected by using the kit is completed by a commercial real-time fluorescent quantitative PCR instrument, the operation is simple, the consumed time is short, and the pollution can be reduced to the maximum extent.

Description

technical field [0001] The invention relates to a kit for detecting Candida glabrata (Candida glabrata), in particular to a kit for detecting Candida glabrata with real-time fluorescent quantitative polymerase chain reaction technology. Background technique [0002] In recent years, with the continuous expansion of immunocompromised populations (including malignant tumors, blood diseases, AIDS, SARS, diabetes, autoimmune diseases, severe burns and trauma, long-term drug addicts, etc.); broad-spectrum antibiotics, corticosteroids and immune Inhibitors are widely used clinically; new technologies such as organ transplantation, catheter intubation, and interventional therapy are widely developed, which has sharply increased the incidence of opportunistic fungal infections and has become one of the main causes of nosocomial infection deaths. [0003] According to data from the US National Nosocomial Infection Surveillance Center (NNIS), the rate of deep fungal infection in hospi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 王前龙燕郑磊
Owner SOUTHERN MEDICAL UNIVERSITY
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