Biological Sample Collection, Transport, Archiving, and Purification for Nucleic Acid Analysis
After sample collection, the other steps do not follow in a specific order. For example, DNA can be prepared from fresh sample, or the sample can first be transported off-site then stored (short term) or archived (long term).
A number of options are available for sample collection, transport, archiving, and DNA purification. These can be categorized into those that are amenable to room temperature archiving and transport, and those that are not. In general, untreated biological samples are not stable at room temperature.
Section Overview
Options that do not allow room temperature archiving and transport
Biological samples collected directly into a tube, multiwell plate, or other vessel are typically not stable at room temperature. To maintain nucleic acid integrity, samples should be processed immediately, stored over the short term (with or without an added chemical stabilizer) at a temperature appropriate for the sample type, or collected onto a matrix suitable for room temperature archiving. For short-term storage, conditions vary by sample type. For example, blood samples can be stored at 4 °C for up to 48 h before isolating DNA, while tissue samples and pelleted cells may be stored at -80 °C for several weeks or months. Cells may be frozen directly, or a stabilizing agent (see below) may be added. Most tissues should be “flash frozen” in liquid nitrogen for best results. Biological samples that require refrigeration or freezing also require wet or dry ice, respectively, for shipping. In addition, human samples must be labeled as potential biohazards. Shipping of these samples to less accessible regions may be limited to certain days of the week, and the shipper must be willing to deal with biohazardous materials.
Chemical stabilizers may be used to stabilize DNA, RNA, and/or protein. These chemicals allow samples to be stored for short periods (several days or weeks) at room temperature or for longer periods (12+ mo) at -20 °C or -70 °C. Chemical stabilizers require an additional kit or method to isolate nucleic acids.
Options that allow room temperature archiving and transport
A straightforward way to prepare a sample for room temperature archiving and transport is to collect the sample directly onto a paper matrix. The options for paper matrices include a variety of untreated matrices (e.g., Guthrie or Whatman 903 cards) and chemically treated matrices (i.e., Whatman FTA technology). When nucleic acids will be isolated, we recommend using chemically treated matrices designed to stabilize nucleic acids and denature proteins. Additional details on Whatman FTA technology are provided later in this chapter.
Other products are available for room temperature archiving. However, these products do not stabilize the nucleic acids in the samples themselves; they stabilize previously purified nucleic acids.
See Table 1 for a summary of the different options for sample collection, transport, archiving, and DNA purification, including advantages and disadvantages.
| Collection | Transport | Can sample be archived? | Additional DNA purification method required? | (+) Advantages/ (−) Disadvantages |
|---|---|---|---|---|
| Use fresh sample | Dry ice for tissues and cells; wet ice for blood Human samples must be treated as biohazardous | No. Sample is used up. | Yes | + Does not require a refrigerator or freezer − Sample must be used right away − Can’t return to sample for additional analysis because sample was not stored |
| Use frozen tissue/cells or use blood stored at 4 °C | Dry ice for tissues and cells; wet ice for blood Human samples must be treated as biohazardous | No. Short-term storage only in freezer or refrigerator, depending on sample type | Yes | + Allows user to return to sample for additional analysis − May require use of liquid nitrogen to freeze tissues − Does not allow for sample archiving |
| Add chemical stabilizers to the sample | Room temperature, wet ice, or dry ice (depends on how long sample is expected to be stable at that temperature) | Yes, in a freezer | Yes | + Inactivates nucleases (and proteases, depending on the chemical) + Samples can be stored for several days or weeks at room temperature − Requires a freezer for long-term storage − Tissue must be cut to be less than 0.5 cm in at least one dimension − Requires an additional DNA purification method |
| Isolate nucleic acids, then stabilize with a commercially available product for this purpose | Room temperature | Yes, at room temperature | Yes. DNA must be prepared before using these archiving methods | + Allows room temperature archiving + Available as single tubes or as 96-well plates |
| Spot sample onto untreated matrix (e.g., Guthrie or 903 card) | Refrigerated to preserve integrity of nucleic acids and proteins Human samples must be treated as biohazardous | Yes, at -20 °C | Yes | + Less expensive than chemically treated paper − Nucleic acids must be purified before using product − Nucleic acids not stabilized at room temperature for extended periods of time − Nucleic acids can be stored at room temperature, but the integrity of the nucleic acids is unknown; amplification of long PCR fragments (> 2 kbp) may not be possible |
| Apply sample onto chemically treated matrix (i.e., Whatman FTA technology) | Room temperature | Yes, at room temperature | No. Requires only FTA Purification Reagent (FTA) or water (FTA Elute) for DNA purification | + Complete system for sample collection, transport, archiving, and DNA purification + Allows room temperature archiving—more than 17 yr (and counting) real-time stability with human blood on FTA + Allows for noninvasive sample collection (i.e., buccal swabs) + Quick and easy DNA purification + Can be used with a variety of sample types from tissue to plant material to cultured cells and bacterial cultures |
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