A classic issue in biology is validation of a new concept on actual human tissue. A frequent problem is there is not enough tissue, either in amount or case number, to complete the analysis. This chronic shortage of tissue is partly due to the fact that the initial sampling is typically done by a pathologist whose primary goal is to provide a diagnostic consultation, not material for future experimentation. The difficulty of working with human tissue is further complicated by increasingly stringent guidelines regarding obtaining informed consent for its use. Thus, when tissue is obtained, it should be optimally managed to maximize its value. Tissue microarray represents a mechanism for highly effective use of this scarce resource.
|Conventional Slides||TMA Advantages|
|• One case per slide||• 50-500+ cases per slide|
|• Requires many batches for a single study||• Entire cohort may be done in a single experiment|
|• Requires large amounts of reagent||• Requires less than 1 ml total reagent volume for entire cohort|
|• Subject to differential antigen retrieval||• Not subject to differential antigen retrieval|
|• Very expensive and slow for large cohorts||• Very rapid for large cohorts|
|• 150 sections of one case yield only 150 assays||• 150 sections with over 500 cases potentially yields up to 75,000 assays|
StainingLike conventional formalin-fixed paraffin embedded material, tissue microarrays are amenable to a wide range of techniques, including histochemical stains, immunologic stains with either chromogenic or fluorescent visualization, in situ hybridization (including both mRNA ISH and FISH), and even tissue micro-dissection and techniques.
Tissue RetentionIn these cases, the block may be cored a few times without destroying the block. Then upon subsequent sectioning, it is still possible to make a diagnosis, even though tissue has been taken for array-based studies.
Are the small histo-spots representative of a whole section?
The major potential limitation of this technique is tissue volume. Skeptics claim that the amount of material analyzed is too small and potentially not representative of the entire tumor. In a reproducibility study, the Sauter group found identical Kaplan Meier curves were generated by analysis of 4 unique sets of spots from the same patients (Torhorst et al, AJP 2001). Thus although any given histo-spot may be negative on a given array, the statistical power of analysis of hundreds or thousands of cases eliminates the affect of variability of a single data point in the ultimate conclusions. Our own study to assess the number of histo-spots required to obtain an equivalent result to a tissue section using the standard breast cancer prognostic markers (Estrogen and progesterone receptors and HER2 oncogene) shows that analysis of only 2 histo-spots results in >95% accuracy (see figure in Camp et al, Lab Invest, 2000). More recently, numerous other studies have found similar results.
We compared the staining of 2 to 10 microarray disks and the whole tissue sections from which they were derived and determined that analysis of two disks is comparable to analysis of a whole tissue section in more than 95% of cases. To evaluate the potential for using archival tissue in such arrays, we created a breast cancer microarray of 8 to 11 cases from each decade beginning in 1932 to the present day and evaluated the antigenicity of these markers and others. This array demonstrates that many proteins retain their antigenicity for more than 60 years, thus validating their study on archival tissues. We conclude that the tissue microarray technique, with 2-fold redundancy, is a valuable and accurate method for analysis of protein expression in large archival cohorts.
The number of sections from an array depends on donor block thickness. Therefore, tissue yield may be limited.
Tissue often does not go completely through the core.