Demonstration of human immunodeficiency virus by colorimetric in situ hybridization: a rapid technique for formalin-fixed paraffin-embedded material.
A colorimetric method of in situ hybridization has been developed for the rapid detection of human immunodeficiency virus (HIV) in formalin-fixed paraffin-embedded material. Following optimization of digestion conditions, biotin-labeled DNA probes are detected with an alkaline phosphatase conjugate. The method is verified using fixed paraffin-embedded cell blocks of HIV-infected and uninfected lymphocyte cell cultures. Hybridization specifically detects both viral RNA and proviral DNA. Formalin fixation for intervals up to 21 d did not significantly hamper the signal under the appropriate digestion conditions; however, Trump's fixation for even 12 h greatly reduced the intensity of the hybridization. This technique for in situ hybridization is amenable to automation, provides results within 6 h, and results in good morphologic preservation. A key feature of the technique is the use of human placental DNA as an endogenous positive control to optimize the empirically determined conditions for protein digestion.
Unger
E R
ER
Department of Pathology and Laboratory Medicine, Emory University, Atlanta, Georgia.
Chandler
F W
FW
Chenggis
M L
ML
Ou
C Y
CY
Warfield
D T
DT
Feorino
P M
PM
D002478:Cells, Cultured; D003124:Colorimetry / Q000379:methods*; D004279:DNA, Viral / Q000032:analysis; D005557:Formaldehyde; D006678:HIV / Q000302:isolation & purification*; D006801:Humans; D008214:Lymphocytes / Q000032:analysis / Q000382:microbiology; D009693:Nucleic Acid Hybridization; D010232:Paraffin; D012367:RNA, Viral / Q000032:analysis
D004279:DNA, Viral; D012367:RNA, Viral; D005557:Formaldehyde; D010232:Paraffin