Long-term preservation of antigenicity on tissue microarrays

Abstract

Tissue microarrays have facilitated the evaluation of large cohort studies; however, there is little data on the best method for preserving sections once they are cut. We assessed three methods of storing precut breast cancer microarray slides: paraffin coating and storage in a nitrogen desiccator, either alone or in combination. We tested the durability of three antigens, cytokeratin, estrogen receptor, and Ki-67 on microarrays stored under these conditions for 3 months at room temperature. Staining was assessed with both manual scoring using traditional brown stain (0-3 þ ) as well as automated scoring using fluorescently stained sections. Staining intensity was compared to that obtained from freshly cut slides. Slides stored under ambient conditions (room temperature and air) for 3 months exhibited marked degradation of all target antigens, in some cases resulting in slides that were virtually unreadable. We found that combined paraffin coating and nitrogen storage resulted in the best preservation of antigenicity, with retention of 72-99% of the antigenicity of a freshly cut slide, depending upon the marker and detection system used. The use of either paraffin coating or nitrogen storage alone protected slides to a lesser degree.

Loss of antigenicity with tissue age in breast cancer

“…To minimize other explanations for antigen loss, we used TMAs and slides cut and stained on the same day to decrease specimen and inter-array variability and to minimize any effects of slide storage, as previously described. 8 While we believe that the effects we have described are a function of tissue aging within the paraffin block, there are limitations to this study. Perhaps most significant is the lack of fresh or very recent tissue from recent months and years.…”

“…8,13,14 Whether these processes affect unsectioned tissue, and to what degree they cause signal loss is not well-described. The data here suggest that signal degradation does occur in this setting.…”

“…After sectioning, significant loss of immunoreactivity in tissue slides has been documented in weeks to months, despite optimal storage conditions including paraffin coating and nitrogen desiccation. 7,8 In contrast to cut and sectioned tissue, protein expression is considered well-maintained in paraffinembedded tissue blocks for many years. 9 Theoretically, unsectioned tissue in paraffin blocks should be less exposed to degrading effects, though the persistence and reliability of antigenicity in these tissues over many years remain poorly characterized.…”

Prognostic Significance of Cadherin-Based Adhesion Molecules in Cutaneous Malignant Melanoma

“…A 0.6 mm tissue microarray was constructed according to published methods (25). Sections (5 Am) were cut from the tissue microarray master using a tissue microtome, transferred to glass slides using a UV crosslinkable tape transfer system (Instrumedics), dipped in paraffin, and stored in a nitrogen chamber to prevent antigen degeneration before staining (26). Immunohistochemistry.…”

Tissue microarrays, tread carefully

“…They also demonstrated the need for redundancy to ensure proper sampling of the tissue. 1 DiVito et al 2 have taken this a step further by showing that prepared TMA slides need to be stored properly in order to prevent the loss of antigenicity.…”