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December 1, 1986Proceedings of the National Academy of Sciences685 citationsOpen Access

Purification and NH2-terminal amino acid sequence of a T-cell-derived lymphokine with growth factor activity for B-cell hybridomas.

JSJacques Van SnickSCSylvie CayphasAVAnne Vink

Key Points

  • The aim was to purify and characterize a T-cell-derived lymphokine that supports the growth of B-cell hybridomas.
  • Used stable factor-dependent B-cell hybridomas for purification monitoring.
  • Employed gel filtration, anion-exchange chromatography, and reversed-phase HPLC for fractionation of the growth factor.
  • Identified the NH2-terminal amino acid sequence of the purified factor.
  • The purified lymphokine showed growth factor activity for B-cell hybridomas at a concentration of 0.1 pM.
  • Molecular mass estimated between 22-29 kDa with variations likely due to glycosylation.
  • The NH2-terminal sequence did not show significant homology with known lymphokines.

Abstract

A T-cell-derived lymphokine was identified by its ability to support the growth of a subset of B-cell hybridomas. Hybrids that failed to survive in the absence of this molecule represented a major proportion of rat-mouse hybridomas but were very rare among mouse-mouse B-cell hybrids. Stable factor-dependent B-cell hybridomas were used to monitor the purification of the growth factor from the supernatant of a clonotypically stimulated mouse helper T-cell clone. Sequential fractionation using gel filtration, anion-exchange chromatography, and reversed-phase HPLC resolved the factor from other B-cell growth factors and yielded a single-chain protein characterized by a major charge (pI = 5-7) and molecular mass (22- to 29-kDa) heterogeneity, probably due to variations in glycosylation. The NH2-terminal amino acid sequence of this protein, which is active on B-cell hybridomas in the 0.1 pM range, showed no significant homology with that of known lymphokines. Because the purified factor also supported the growth and survival in vitro of murine plasmacytomas (to be published elsewhere), it was provisionally designated interleukin-HP1 (where H stands for hybridoma and P stands for plasmacytoma).

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Cite This Study

Snick et al. (1986) studied this question.

synapsesocial.com/papers/6a153f6fd64fa333899f6c6ahttps://doi.org/10.1073/pnas.83.24.9679
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