Immunohistochemical staining of plastic embedded bone marrow trephine biopsy specimens after microwave heating.

Abstract

Bone marrow trephine biopsies are routinely performed, in conjunction with marrow aspirates, in the investigation of patients with a variety of haemopoietic disorders, both neoplastic and non-neoplastic. Trephine biopsy specimens are particularly useful where there is significant bone marrow fibrosis which frequently results in an inadequately aspirated specimen or "dry tap". Trephine biopsy specimens are also useful in the assessment of marrow cellularity and in the determination of the extent of marrow involvement by neoplastic or other infiltrates. The relation between cellular elements and normal marrow structures, such as bony trabeculae and blood vessels, can be assessed accurately only on bone marrow trephine biopsy specimens. In addition focal lesions, such as granulomas, are more readily identifiable on trephine biopsy specimens than on marrow aspirates.Two main methods are used for the routine histological examination of bone marrow trephine biopsy specimens, namely paraffin wax embedding followed by decalcification and plastic embedding followed by the cutting of 1 ,um sections. A third approach to processing such specimens is the use of cryostat sections of undecalcified bone marrow biopsy specimens. This method, although allowing for optimal antigen preservation, results in very poor cellular morphology, a serious obstacle in evaluating these specimens, and thus has not gained widespread acceptance. Problems arise in cutting sections of bone marrow trephine biopsy specimens because of the intimate mixture of hard tissue (bone) and soft tissue (marrow and fat). To cut adequate intact sections, one can either make the tissue uniformly soft by paraffin wax embedding followed by decalcification, or make the tissue uniformly hard by using a plastic embedding procedure. There has been much debate in the literature regarding the relative merits of these two techniques.`'7 Those who advocate paraffin wax embedding of trephine biopsy specimens stress the advantages of the familiarity of most pathologists with sections cut from such material combined with their unfamiliarity with plastic embedded sections and the fact that widespread antigen preservation allows a wide range of immunohistochemical reactions to be performed.'2 Furthermore, plastic embedding represents a significant departure from normal routine for the diagnostic histopathology laboratory as special equipment, different preparation protocols

Effect of Bone Decalcification Procedures on DNA In Situ Hybridization and Comparative Genomic Hybridization: EDTA Is Highly Preferable to a Routinely Used Acid Decalcifier

“…Moreover, antigenic sites, DNA, and mRNA are preserved. Recent developments in plastic technology, especially the introduction of new resins that can be removed after sectioning, as well as new methods for antigen retrieval, indicate that DNA and mRNA ISH on plastic-embedded tissue will become more feasible in the near future (McCluggage et al 1995;Church et al 1997;Warren et al 1998). …”

Optimal processing of bone marrow trephine biopsy: the Hammersmith Protocol

“…2 6 Several papers deal with these issues and confirm adequate antigen and DNA preservation in resinembedded BMT specimens. [7][8][9][10] However, RNA preservation and suitability for ISH analyses of mRNA is not clear. The main criticism of this procedure is the requirement of specialised technology and skill and the additional costs associated with it.…”

How we process trephine biopsy specimens: epoxy resin embedded bone marrow biopsies

“…11 13 Alternatively, resin protocols can be adjusted to the requirements of antigen and even DNA detection. [25][26][27][28] We have run parallel resin and paraffin wax embedding for bone marrow trephine biopsy specimens. Paraffin wax embedded specimens may be used for consulting with centres not familiar with resin technology and for routine DNA extraction.…”

“…34 In addition, aliphatic epoxy resins such as those based on the Epon 812 formula can be used for immunohistochemistry in a similar way to that described here. 27 Before the era of heat induce antigen retrieval, we exploited the etching effect of sodium methoxide on epoxy resins (thought to be the result of transesterification and/or oxidation 29 ) for retrieving antigens and enhancing the sensitivity of immunodetection in paraffin wax embedded sections. 35 Recently, this approach has been advocated as a non-heat mediated antigen retrieval technique.…”