Scope of document This UK SMI describes key elements of how to organise facilities for molecular amplification assays. This includes designated rooms, workflow plan, reagents, consumables and staff within a molecular diagnostic laboratory1. This UK SMI should be used in conjunction with other UK SMIs. Introduction The ability of polymerase chain reaction (PCR) to produce large numbers of copies of a target sequence from minute quantities - sometimes single copies of DNA has provided the exquisite sensitivity that makes PCR a powerful diagnostic tool. However, this ability also necessitates that extreme care needs to be taken to avoid the generation of false positive results. False positive results can occur from sample-to-sample contamination and from the carry-over of amplicons from a previous amplification of the same target. Another significant risk is through cross-contamination of different reactions prepared at the same time and contamination of master stocks (for example oligonucleotide stocks) by DNA templates which is a major risk to be considered2. In addition to cloned DNA and virus cell cultures, microorganisms within the environment are a significant source of contamination. This can be limited by meticulous attention to good laboratory procedures3. Thought should be given to facility design and operation within clinical laboratories in which polymerase chain reactions are performed. Laboratories should be designed to prevent exposure of pre-PCR reagents and materials to PCR products/contaminants. Consideration needs to be given to other activities that could cause contamination such as shared specimens between laboratories as well as handling of “cultured” samples in molecular laboratory workspace. Cultured isolates present a contamination risk that could compromise primary molecular diagnostic assays from primary samples. This document describes procedures that will help to minimise the carry-over of amplified nucleic acid.
Scope of document This UK SMI describes key elements of how to organise facilities for molecular amplification assays. This includes designated rooms, workflow plan, reagents, consumables and staff within a molecular diagnostic laboratory1. This UK SMI should be used in conjunction with other UK SMIs. Introduction The ability of polymerase chain reaction (PCR) to produce large numbers of copies of a target sequence from minute quantities - sometimes single copies of DNA has provided the exquisite sensitivity that makes PCR a powerful diagnostic tool. However, this ability also necessitates that extreme care needs to be taken to avoid the generation of false positive results. False positive results can occur from sample-to-sample contamination and from the carry-over of amplicons from a previous amplification of the same target. Another significant risk is through cross-contamination of different reactions prepared at the same time and contamination of master stocks (for example oligonucleotide stocks) by DNA templates which is a major risk to be considered2. In addition to cloned DNA and virus cell cultures, microorganisms within the environment are a significant source of contamination. This can be limited by meticulous attention to good laboratory procedures3. Thought should be given to facility design and operation within clinical laboratories in which polymerase chain reactions are performed. Laboratories should be designed to prevent exposure of pre-PCR reagents and materials to PCR products/contaminants. Consideration needs to be given to other activities that could cause contamination such as shared specimens between laboratories as well as handling of “cultured” samples in molecular laboratory workspace. Cultured isolates present a contamination risk that could compromise primary molecular diagnostic assays from primary samples. This document describes procedures that will help to minimise the carry-over of amplified nucleic acid.
Q4i5.docx