Combination of Adhesive-tape-based Sampling and Fluorescence <em>in situ</em> Hybridization for Rapid Detection of <em>Salmonella</em> on Fresh Produce

Bledar Bisha; Byron F. Brehm-Stecher

doi:10.3791/2308

JoVE Journal
Immunology and Infection

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JoVE Journal Immunology and Infection

Combination of Adhesive-tape-based Sampling and Fluorescence in situ Hybridization for Rapid Detection of Salmonella on Fresh Produce

胶粘剂基于磁带的采样和荧光组合原位杂交快速检测沙门氏菌新鲜农产品

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09:10 min

October 18, 2010

DOI: 10.3791/2308-v

Bledar Bisha¹, Byron F. Brehm-Stecher²

¹Center for Meat Safety and Quality, Department of Animal Sciences,Colorado State University, ²Rapid Microbial Detection and Control Laboratory, Department of Food Science and Human Nutrition,Iowa State University

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Overview

This protocol describes a method for sampling the surfaces of fresh produce, such as tomatoes, using adhesive tape. It enables rapid detection of Salmonella through fluorescence in situ hybridization (FISH).

Key Study Components

Area of Science

Microbiology
Food Safety
Molecular Biology

Background

Salmonella is a significant foodborne pathogen.
Traditional methods for detecting Salmonella can be time-consuming.
Fluorescence in situ hybridization (FISH) allows for rapid detection.
Adhesive tape sampling is a novel approach for surface sampling.

Purpose of Study

To develop a simple method for sampling fresh produce surfaces.
To enable rapid molecular detection of Salmonella.
To improve food safety protocols in produce handling.

Methods Used

Application of adhesive tape to collect surface-bound cells.
Cell fixation and dehydration in an ethanol series.
Hybridization using a Salmonella-specific oligonucleotide probe cocktail.
Washing and analysis of samples via fluorescence microscopy.

Main Results

Successful extraction of Salmonella from fresh produce surfaces.
Rapid detection achieved through FISH methodology.
Demonstrated effectiveness of adhesive tape sampling.
Potential for enhancing food safety measures.

Conclusions

The adhesive tape method is a viable option for sampling.
FISH provides a rapid detection technique for pathogens.
This approach can be integrated into food safety practices.

Frequently Asked Questions

What is the main advantage of using adhesive tape for sampling?

Adhesive tape allows for easy collection of surface-bound cells without complex equipment.

How does FISH work in detecting Salmonella?

FISH uses fluorescent probes that bind specifically to Salmonella RNA, allowing for visualization under a microscope.

Can this method be applied to other types of produce?

Yes, the adhesive tape sampling method can be adapted for various fresh produce surfaces.

What are the limitations of this method?

Potential limitations include the need for careful handling to avoid contamination and the specificity of the probes used.

Is prior knowledge of microbiology required to perform this protocol?

Basic understanding of microbiological techniques and safety protocols is recommended.

How long does the entire process take?

The entire sampling and detection process can be completed in a few hours, making it relatively quick.

本协议描述了简单的基于磁带的粘合剂为番茄等新鲜农产品表面取样的方法，全细胞快速检测

该程序的总体目标是将基于胶带的新鲜农产品采样与使用 C 2 杂交（也称为鱼）中的快速荧光对沙门氏菌物种进行全细胞分子检测相结合。这是通过首先将胶带贴在所需材料上并提取表面结合的细胞来实现的。电池充电带可以直接分析，也可以进行固相或液相富集。

该程序的第二步是在胶带上，对于在肉汤中暗示或富集在肉汤中的细胞，在乙醇系列中进行自固定和脱水。第二步涉及将细胞固定在悬浮液中，然后将细胞悬浮在储存缓冲液中。该程序的第三步是使用沙门氏菌特异性寡核苷酸探针混合物进行快速磁带杂交步骤，然后用洗涤缓冲液冲洗去除多余的探针（对于通过传真分析的样品），鱼步骤在悬浮液中进行，并且可以在分析前使用简单的稀释步骤去除多余的探针。

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