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.
Department of Pathology and Laboratory Medicine, Emory University, Atlanta, Georgia.
E R
ER
Unger
F W
FW
Chandler
M L
ML
Chenggis
C Y
CY
Ou
D T
DT
Warfield
P M
PM
Feorino
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