Isolation of human immunodeficiency virus from peripheral blood lymphocytes stored in various transport media and frozen at -60 degrees C.
The peripheral blood lymphocytes from 48 heparinized blood specimens from human immunodeficiency virus (HIV) antibody-positive individuals were divided into two aliquots. One aliquot was reconstituted in one of the following five media. Medium 1 consisted of tryptose broth with 0.5% gelatin; medium 2 consisted of RPMI 1640 containing 10% fetal bovine serum (FBS); medium 3 consisted of RPMI 1640 containing 20% FBS, Polybrene, interleukin 2, and anti-alpha interferon; medium 4 consisted of medium 2 plus 10% dimethyl sulfoxide (DMSO); and medium 5 consisted of medium 3 plus 10% DMSO. Lymphocytes were stored in these five media at -60 degrees C. The other aliquot of cells was stored at -190 degrees C in RPMI 1640 containing 50% FBS and 10% DMSO. After 1 week, both aliquots were cocultivated with phytohemagglutinin-stimulated uninfected peripheral blood lymphocytes, and presence of HIV was detected by the reverse transcriptase test. Storage in medium 1, 2, or 3 did not result in satisfactory isolation rates, but storage at -60 degrees C in medium 4 or 5 gave equal or better isolation rates than did storage at -190 degrees C. Inactivation of HIV by freezing of the cells without DMSO correlated with high antibody titers to core and polymerase proteins as measured by Western (immuno-) blotting.
Gallo
D
D
Viral and Rickettsial Disease Laboratory, State of California Department of Health Services, Berkeley 94704.
Kimpton
J S
JS
Johnson
P J
PJ
D000163:Acquired Immunodeficiency Syndrome / Q000097:blood* / Q000382:microbiology; D001793:Blood Preservation; D015153:Blotting, Western; D003470:Culture Media; D004121:Dimethyl Sulfoxide; D005615:Freezing; D006678:HIV / Q000254:growth & development / Q000302:isolation & purification*; D006801:Humans; D008214:Lymphocytes / Q000382:microbiology*
D003470:Culture Media; D004121:Dimethyl Sulfoxide