It is often said that the best place to catch a disease is in a hospital; likewise, the most common place for sample contamination to occur is in the analytical laboratory. Sample contamination may be defined as the inadvertent addition of target or detectable analytes to samples during the sample collection, transportation, preparation, or analysis processes. Consequently, virtually all laboratories have contamination problems and these problems are, to a large extent, unavoidable and insurmountable without strict attention to procedural validation, monitoring, and routine instrument maintenance.
Common sources of contamination include organic or inorganic analytes in atmospheric dust and sample residues. Sample residues are left behind on improperly cleaned tools such as pipettes, syringes, spatulas, as well as storage or volumetric containers. A clean sample in contact with a contaminated surface is by definition a contaminated sample. In some cases, careless or improper sampling techniques which expose the sample to air, fluid, or surface borne analytes are also a source of contamination. In these instances, proper training of personnel and the implementation of validated cleaning, handling, and storage procedures can help reduce contamination-related errors while saving money, effort, and time.
Sample preparation or pretreatment procedures are another common source of contamination. Solvents used in extraction or purification procedures or the addition of diluents, buffers, or analytical reagents may introduce foreign or unintended analytes that may interfere with the final analysis or lead to incorrect or misleading conclusions. Proper raw material and lab supply intake procedures can help ensure that contamination issues were not the result of a laboratory equipment supplier.
Far less obvious sources of contamination are the analytical standards and blanks which are an integral and indispensable component of virtually all analytical procedures. Many analysts automatically assume that analytical standards or reagents prepared as blanks are accurate and free of contaminants, leading to a potentially false conclusion that the source of any analytes detected is the sample. Here again, validation, procedural clarity, monitoring, and training are the only reliable defense.
In many cases, the removal of excess extraction or reaction solvent is a necessary precursor to the analytical process. There are a number of accepted methods for solvent removal and sample concentration including rotary evaporation, distillation, oven drying, freeze drying, and gas assisted evaporation, i.e., “nitrogen blow down”. Each of these methods is a potential source of contamination so care must be exercised in the proper application of these techniques.
When gas assisted evaporation systems such as Organomation’s N-EVAPs, MULTIVAPs or MICROVAPs are utilized for sample preparation and the removal of excess solvent, it is always advisable to:
1. Make sure that sample vials are properly and verifiably cleaned prior to the introduction of the sample or sample containing solution. Disposable, single use sample vials are the preferred choice but if reusable sample vials are used they should be cleaned and stored following a validated cleaning and storage procedure.
2. Make sure that gas delivery needles are clear and free of any residues. Ideally needles should be cleaned before each application and examined before use.
3. Make sure the gas source is clean and dry. If bottled nitrogen or other inert gas is being used the gas should be high purity (99.99%) or ultra high purity (99.999%) in situations where higher precision is desired. In cases where the sample is moisture sensitive or susceptible to microbial attack, the gas should be passed through a dryer and a micro filter. If atmospheric air (usually from a compressor) is used, the gas must be passed through a dryer and a micro filter as well.
4. Take care when mounting sample vials in the nitrogen evaporator. Do not allow the sample or sample containing solution to come into contact with the needle. Before mounting the sample vial raise the needle high enough to allow the sample vial to be seated in the holder without tipping and then lower the needle into position after the sample is in position.
5. Avoid creating excessive sample turbulence inadvertently. Set the gas flow rate at a lower setting initially and then increase the flow using the needle valve. If a series of samples are being processed this is only necessary for the initial series and the setting may be used for subsequent samples provided the same sample vials and solution volumes are maintained. Splashing or spattering may result in sample loss, cross contamination, and needle contamination or soiling.
6. Make sure that the bath temperature is sufficiently high to produce a satisfactory evaporation rate. However, too high of a temperature will create condensation on the exposed surfaces of the instrument. Condensed moisture or solvent may drip into the sample or leave residues on the needle or surfaces above the sample.
7. Take care when removing sample vials from the instrument; raise the needles as high as possible to ensure that the sample vial can be removed without contact with the needle. The needle must not come into contact with the sample vial walls or the sample in the vial. It is also advisable to discontinue the gas flow due to the possibility that by lifting the tube vertically out of the vial holder the sample could be brought into near contact with the needle which could result in turbulent splashing or spattering.
Again, all laboratories have contamination problems that can only be controlled by the implementation of an active program designed to continuously monitor the many different types and sources of contamination. Reviewing, validating, and training personnel on these preventive measures will ultimately save the laboratory both time and money.
INFORMATION RELATED TO cGLP, TRAINING & REGULATIONS
1. Barge, Maureen S. and Ussar, James P., Good Laboratory Practice Standards: Applications for Field and Laboratory Studies (ACS Professional Reference Books), American Chemical Society, 1992
2. Seller, Jurg P., Good Laboratory Practice: The Why and The How, 2005
3. Allport – Settle, Mindy J., Good Laboratory Practice: Nonclinical Laboratory Studies Concise Reference
4. Huber, Ludwig, Validation and Qualification in Analytical Laboratories, 2nd Edition, 2007
5. Huber, Ludwig, Good Laboratory Practice and Current Good Manufacturing Practice, Agilent Technologies Deutschland Gmbh, 2002
6. Compliance and Safety Training, Preventing Contamination in the Laboratory, DVD Program (http://complianceandsafety.com)
Additional sources of information regarding cGLP, training and regulations applicable to specific industries or field are available from:
-Federal Registry: 21 CFR 211, Subpart I (4)
-American Chemical Society
-ASQ (American Society for Quality)
-American Association for Laboratory Accreditation
-Compliance and Safety Trainining (http://complianceandsafety.com)