An Experience From the Cooperative Human Tissue Network
The scientific usefulness of the data obtained from tissue analysis is related to specimen quality, which may be affected by conditions that may contribute to the degradation of the specimen before processing and analysis.

We determined the usability of nucleic acids extracted from banked human tissues for further molecular analyses. We assayed 151 tissue specimens, stored for various times at 4 divisions of the Cooperative Human Tissue Network, National Cancer Institute, Bethesda, MD, for DNA and RNA degradation. Simple electrophoresis, polymerase chain reaction (PCR), reverse-transcriptase (RT)-PCR, and Northern blot analysis were compared to determine the optimal quality control procedure. In addition, a time course degradation procedure was performed on human lung tissue.

Gel electrophoresis was as informative as PCR, RT-PCR, and Northern blot analysis in determining the molecular usefulness of the human tissues. Overall, 80% of the stored human tissues had good-quality DNA, and 60% had good-quality RNA. Electrophoresis procedures for DNA and RNA offer a quick and valuable measure of the molecular quality of stored human tissues. The DNA and RNA degradation of one tissue type (lung) was stable for both nucleic acids for up to 5 hours after excision.

Clinical and molecular pathology are undergoing a revolution based on the accelerated advances in biotechnology such as DNA microarrays and proteomics. Answers to fundamental questions such as "How does the DNA sequence differ between individuals?" and "What makes one individual more prone for a certain disease?" are eagerly sought in this postgenomic era. Several novel genes and their products have been identified in human cancers by screening archival tissue samples using molecular methods. More important, molecular diagnostic testing is requested more often in certain clinical conditions, eg, testing for T- or B-cell clonality in early cutaneous lymphomas.

The scientific usefulness of the data obtained from the analysis of tissues is related directly to the quality of the tissue specimen. The most ideal tissue specimen is one that carries a complete and unaltered representation of the tissue in vivo. While this may be close to impossible ex vivo, it is critical that we understand and attempt to improve on the quality of research specimens. Unlike animal tissues, which can be controlled to maximize the quality of the specimen, human tissues used for research often are subjected to conditions that may contribute to the degradation of the specimen before they can be appropriately processed and analyzed. For example, factors such as the time the vascular system is compromised before surgical removal of a tissue may affect the quality of the tissue. With the growing number of investigators requesting human tissues for research and an increasing use of tissue microarrays and high-throughput molecular profiling to evaluate diseased tissue, it is important to provide investigators with information about the molecular quality of the tissue.

An often-used method for rapid evaluation of the yield of the nucleic acid extracted from cells or tissue is to determine the ratio of optical absorbance at 260 and 280 nm. However, the ratio suggests degree of protein contamination and does not necessarily reflect the quality of nucleic acid. The expression of 28S ribosomal RNA (rRNA) expression is considered the best marker for assessing the integrity of the nucleic acid in the tissue because it remains stable in stimulated and nonstimulated conditions. Several factors, including the specimen type, preexcision hypoxia, preservation treatment of the tissue, extraction method, type and length of storage, and freeze and thaw factors affect the molecular quality of the tissue.

General dogma suggests the following: (1) Acellular and pancellular tissues are less labile than densely cellular tissue. (2) Hypoxic conditions initiate cell death mechanisms and subsequent degradation. (3) Quick treatment to inactivate degrading enzymes is important for preserving tissue and nucleic acid integrity. (4) Fresh, unfixed tissue is better than chemically fixed tissue for extracting nucleic acids. (5) Short periods of storage yield better quantity and quality of nucleic acid than longer periods. (6) With frozen tissue, nucleic acid preservation is enhanced by thawing the tissue as quickly as possible.

Human tissues procured for research are frozen and stored for variable periods. Conventionally, it is thought that the longer the period of storage, the lower the yield or quality of the nucleic acid. Preliminary data from a study at the University of Alabama at Birmingham (W.H.G., oral communication, November 2000) and a recent survey by us (unpublished data) suggested that the vast majority of the investigators rated tissues procured from the Cooperative Human Tissue Network (CHTN) as a good source of DNA and RNA usable for further molecular analysis. However, there is a lack of a controlled study to validate these observations. The purpose of the present study was to determine the usability of nucleic acids extracted from banked human tissues for further molecular analyses. We suggest a quality control method for evaluating the molecular integrity of the tissue before distribution.