From:
http://www.nfstc.org/pdi/Subject04/pdi_s04_m04.htm
Contamination
Author: Debbie Figarelli
Debbie Figarelli serves as DNA Technical Leader at the National Forensic Science Technology Center. Debbie assists with the development of DNA training programs and participates in compliance audits of DNA laboratories.
Current methods used by forensic DNA laboratories are sensitive and capable of amplifying and detecting low quantities of DNA. With the ability to detect minute quantities of DNA, comes an increased chance of detecting contaminant DNA in both samples and controls. Contamination can happen during any step of the process and can result from a variety of substances, including plant material, bacteria, and human genomic DNA. There are occasions when the source of contamination cannot be determined and/or the incidences are sporadic in nature.
It is not always possible to prevent contamination; however, laboratories should ensure that procedures are in place to:
Minimize the risk of contamination
Detect contamination
Document and implement corrective measures for incidents of contamination
This module focuses on extraneous human genomic DNA deposited after the crime event (during the collection and/or analysis process).
The most common sources of extraneous DNA into the collection and analysis processes are: 01-04
Investigators and laboratory staff
Reagents and consumables
Sample cross contamination during analysis
Sample Cross-Contamination
Many laboratories process samples in batches to streamline analyses. One risk of batch analysis is the inadvertent cross-contamination of DNA from one sample to another sample that was processed concurrently. Most detected sample-to-sample sample contamination will be from samples with higher concentrations of DNA to those with lower concentration.
Reference and Evidentiary Sample Processing
One concern with contamination is that an individual may be falsely linked to a crime. Reference samples are generally good quality DNA samples and result in high quantities of extracted DNA. Many laboratories process samples in a way that isolates evidentiary samples from reference samples during the screening, extraction, and PCR stages. Therefore, the possibility of contaminating an evidentiary sample with reference DNA is avoided.
Detection
Contamination introduced in the laboratory analysis process may not be avoidable, making implementation of procedures to detect these incidences crucial. DNA profiles derived from evidentiary samples and reference samples are uploaded into CODIS and are used to associate individuals with a crime. It is imperative that laboratory procedures are in place to ensure the integrity of the data generated for these functions.
Most manufacturer's multiplexes demonstrate optimum efficiency when approximately 1ng of template DNA is analyzed for 28-30 amplification cycles.06 However, the lower limit of detection can be less than 250 pg. Most contamination events involve small quantities of DNA and therefore will be detected at lower threshold values. Laboratories establish reporting thresholds based on their validation studies. Because most contamination is below that threshold, it will not be reported; analysts should assess any allelic activity under the reporting threshold to determine if it could be from contamination.
Some additional detection processes include:
Assessment of controls
Review of batch profiles
Maintenance of unsourced contamination profiles
Establishment of reference databases
Assessment of Controls
As stated above, negative controls and reagent blanks can greatly assist in the detection of contamination. Positive controls and samples from known sources may also aid in the detection of contamination. Positive controls are single source samples of a known type; the detection of additional alleles may indicate contamination. Reference samples are expected to be from a single-source, and while the DNA profile may be unknown, results that indicate a mixture could be a sign of contamination.
