PCR-in situ hybridization detection of human T-cell lymphotropic virus type 1 (HTLV-1) tax proviral DNA in peripheral blood lymphocytes of patients with HTLV-1-associated neurologic disease.
PCR-in situ hybridization (PCR-ISH) was developed and utilized to determine the distribution of human T-cell lymphotropic virus type 1 (HTLV-1) tax proviral DNA in peripheral blood lymphocytes (PBL) from patients with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). PCR-ISH of HTLV-1 tax DNA in PBL from patients with HAM/TSP revealed that 1 in 5,000 to 1 in 10,000 PBL contained virus. PCR-ISH was sensitive, because a positive signal was consistently demonstrated from the HTLV-1-infected cell lines HUT-102 (which contains four to six copies of HTLV-1 proviral DNA per cell) and MT-1 (which contains one to three copies of HTLV-1 proviral DNA per cell). Also, intracellular amplification by PCR-ISH significantly increased sensitivity compared with conventional ISH and was shown to be specific for HTLV-1 tax DNA. These results are in contrast to solution-phase PCR amplification in which greater than 1% of cells were estimated to be infected. The discordance between these results is discussed and may indicate that more than one copy of HTLV-1 tax proviral DNA is present in an individual PBL.
Levin
M C
MC
Neuroimmunology Branch, National Institute of Neurological Diseases and Stroke, NIH, Bethesda, MD 20852, USA.
Fox
R J
RJ
Lehky
T
T
Walter
M
M
Fox
C H
CH
Flerlage
N
N
Bamford
R
R
Jacobson
S
S
D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification*; D016356:Gene Products, tax / Q000235:genetics*; D015368:Human T-lymphotropic virus 1 / Q000302:isolation & purification*; D006801:Humans; D017403:In Situ Hybridization; D008214:Lymphocytes / Q000821:virology; D008969:Molecular Sequence Data; D015493:Paraparesis, Tropical Spastic / Q000097:blood / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D011533:Proviruses / Q000235:genetics / Q000302:isolation & purification; D012680:Sensitivity and Specificity
D017931:DNA Primers; D004279:DNA, Viral; D016356:Gene Products, tax