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Gary S. Wood, MD; Alice Ruffo, MS; Anupama Salvekar, MS; Will Henghold, MD; Morishige Takeshita, MD; Masahiro Kikuchi, MD

Arch Dermatol. 1996;132(11):1339-1343.

Abstract
Objective and Design
To develop a method for the detection of human T-cell lymphotropic virus type I (HTLV-I) in archival biopsy specimens. A polymerase chain reaction—based gene amplification method was developed to detect HTLV-I proviral DNA in paraffinembedded specimens. The specificity of the polymerase chain reaction products was controlled by Southern blot analysis using a nested oligonucleotide probe and by nucleotide sequencing. The nucleophosmin gene and the T-cell receptor- gene were used as controls for the integrity and adequacy of total DNA and T-cell DNA, respectively. This study was conducted with patients referred to an academic medical center. Biopsy specimens were obtained from lesional skin or lymph node from Japanese patients with HTLV-I seropositive adult T-cell leukemia/lymphoma. The main outcome measure was the ability to detect HTLV-I pX region proviral DNA.

Results
Comparative analysis of DNA extracted from fresh samples of the HTLV-I infected MT4 T-cell line demonstrated that formalin fixation and paraffin embedding resulted in a 100-fold reduction in sensitivity of the assay. Nevertheless, HTLV-I pX sequences were still readily detectable in paraffin-embedded samples of MT4 T cells and adult T-cell leukemia/lymphoma specimens. Both formalin and B5 fixation were suitable for the assay that was 100% specific for HTLV-I—infected tissues.

Conclusions
The use of this method should greatly facilitate investigation of the role of HTLV-I in human diseases by allowing analysis of a wide variety of archival tissue specimens. In addition, the controls designed for the current study can be used in a variety of other polymerase chain reaction—based studies of T cells to ensure against false-negative results caused by DNA degradation or inadequate T-cell density.

Arch Dermatol. 1996;132:1339-1343

Author Affiliations
From the Departments of Dermatology and Pathology, and the Skin Diseases Research Center, Case Western Reserve University, and the VA Medical Center, Cleveland, Ohio (Drs Wood and Henghold and Mss Ruffo and Salvekar); and the Department of Pathology, Fukuoka University, Fukuoka, Japan (Drs Takeshita and Kikuchi).

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