Abstract
We have purified to homogeneity the α L-component (70,000–90,000) of the human LT cytolytic system. This lymphokine was purified ~ 10, 000-fold from supernatants of lectin-stimulated human tonsil and adenoid lymphocytes by molecular sieving, ion-exchange chromatography on DEAE-Sepharose, and preparative PAGE. The homogeneity of the radiolabeled molecule was confirmed both by electrophoresis and electrofocusing. The identity of the labeled peak with the lytic activity was demonstrated with a concomitant bioassay of the electrophoresed preparation; in addition, immunoprecipitation with a heterologous specific anti-α L antiserum showed simultaneous precipitation of the radiolabeled component and lytic activity. Immunological and biochemical evidence has previously shown this molecule to be a subunit of the Cx- and α H-LT forms. The latter LT classes are of intense interest because of their capacity for rapid selective cell lysis.
Highlights
LT was originally described as a weak, nonspecific growth-inhibitory or cell-lytic lymphokine (Ruddle & Waksman, 1968; Granger & Williams, 1968; Jeffes & Granger, 1975)
The methods involved in the preparation of human lymphocyte supernatants, in generating the various mol. wt LT classes by molecular sieving of the supernatants, and in resolution of the or,LT class into its charge subclasses by ionexchange chromatography have been published previously in detail (Granger et al, 1978; Lewis et al, 1976) and will only be briefly summarized here
Radioactive fractions containing lytic activity from analytical PAGE were subjected to isoelectricfocusing in tube gels
Summary
LT was originally described as a weak, nonspecific growth-inhibitory or cell-lytic lymphokine (Ruddle & Waksman, 1968; Granger & Williams, 1968; Jeffes & Granger, 1975). Wt forms (alpha-light, 70,000-90,000 d; beta, 35,00&50,000 d; gamma, 12,0~20,000 d) are capable only of protracted lysis of a limited number of continuous cell lines; the larger forms (complex, > 200,000 d; alpha-heavy, 120,00~160,000 d) are capable of causing rapid nonspecific lysis of a spectrum of allogeneic cells, and selective lysis when obtained from lectinstimulated alloimmune immune lymphocytes (Yamamoto et al, 1979; Hiserodt et al, 1978a) Both immunological and biochemical evidence suggests that these classes are comprised of related subunits (Yamamoto et al, 1978; Hiserodt et al, 19786). Subclass, an important subunit of the Complex and alpha-heavy classes
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