RAPID AUTOMATED COMBINED IN-SITU HYBRIDIZATION AND IMMUNOHISTOCHEMISTRY FOR SENSITIVE DETECTION OF CYTOMEGALOVIRUS IN PARAFFIN-EMBEDDED TISSUE BIOPSIES

The authors questioned whether an automated kinetic mode assay of comb ined cytomegalovirus (CMV) late viral message and immediate and early antigens might result in a more sensitive and timely CMV diagnosis rel evant to speedy treatment in the transplant setting. Toward this end, two cohorts were studied using automated in situ hybridization (ISH) f or CMV as well as immunohistochemistry (IHC). The first cohort of pati ents consisted of 19 cases that were histologically positive (CR IV-as sociated cytopathic change). A second cohort consisted of 10 cases tha t were histologically negative, yet culture positive. From the first c ohort of histologically positive cases, 100% were positive by both ISH and IHC run on separate slides. In the second cohort, CMV was detecte d overall in 70% of cases (50% by ISH alone and 30% by IHC alone). The se results indicate that a combined assay of ISH and IHC can detect mo re cases than routine hematoxylin and eosin staining or either assay a lone. In two illustrative cases, used to demonstrate the feasibility o f combining ISH and IHC, the authors used a combined two-color assay ( ISH and IHC) performed sequentially on the same slide. The combined as says resulted in colocalized single cell message and protein in some c ells and demonstrated more positive cells overall (some positive by IH C alone, some by ISH alone, and some by both) than either assay alone, The combined dual color assay can be completed within 4 to 5 hours gi ving the prospect of a same day result, which is faster than shell via l technique with immunofluorescence (24 to 48 hours) or culture (7 to 14 days). This study demonstrates that combining CMV message and prote in assays results in a more sensitive assay and, when carried out in t he kinetic mode, allows a speedy result relevant to early anti-CMV the rapy.