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JMD 2006, Vol. 8, No. 2
Copyright © 2006 American Society for Investigative Pathology & Association for Molecular Pathology

Technical Advance

RCL2, a New Fixative, Preserves Morphology and Nucleic Acid Integrity in Paraffin-Embedded Breast Carcinoma and Microdissected Breast Tumor Cells

Christophe Delfour^*,, Pascal Roger^*, Caroline Bret, Marie-Laurence Berthe, Philippe Rochaix^¶, Nicolas Kalfa^||, Pierre Raynaud^*, Frédéric Bibeau, Thierry Maudelonde and Nathalie Boulle

From the Laboratoires d’Anatomie Pathologique, * Biologie Cellulaire, and Hormonologie, ^|| Centre Hospitalier Universitaire de Montpellier, Hôpital Arnaud de Villeneuve, Montpellier; INSERM U540, Montpellier; the Laboratoire d’Anatomie Pathologique, Centre de Recherche et de Lutte Contre le Cancer Val d’Aurelle, Montpellier; and the Laboratoire d’Anatomie Pathologique, ^¶ Centre de Recherche et de Lutte Contre le Cancer Claudius Régaud, Toulouse, France

Abstract

Methacarn and RCL2, a new noncrosslinking fixative, were compared to formalin-fixed or frozen tissue samples of the same invasive breast carcinoma and were evaluated for their effects on tissue morphology and immunohistochemistry as well as DNA and RNA integrity. The histomorphology of methacarn- or RCL2-fixed paraffin-embedded tumors was similar to that observed with the matched formalin-fixed tissues. Immunohistochemistry using various antibodies showed comparable results with either fixative, leading to accurate breast tumor diagnosis and determination of estrogen and progesterone receptors, and HER2 status. Methacarn and RCL2 fixation preserved DNA integrity as demonstrated by successful amplification and sequencing of large DNA amplicons. Similarly, high-quality RNA could be extracted from methacarn- or RCL2-fixed paraffin-embedded MCF-7 cells, whole breast tumor tissues, or microdissected breast tumor cells, as assessed by electropherogram profiles and real-time reverse transcriptase-polymerase chain reaction quantification of various genes. Moreover, tissue morphology and RNA integrity were preserved after 8 months of storage. Altogether, these results indicate that methacarn, as previously shown, and RCL2, a promising new fixative, have great potential for performing both morphological and molecular analyses on the same fixed tissue sample, even after laser-capture microdissection, and can open new doors for investigating small target lesions such as premalignant breast lesions.