Table I.

L and H chain usage in 3H9.KI hybridomas after cGVH inductiona (38 clones)

Anti-dsDNA+b (n = 26 (68.5%))Anti-dsDNA (n = 12) (31.5%))
3H9tg+c (n = 17 (65%))d3H9tg (n = 9 (35%))3H9tg+ (n = 3 (25%))3H9tg (n = 9 (75%))d
L chain usagee
J11 (6%)1 (11%)04 (44%)
J23 (17%)2 (22%)1 (33%)0
J401 (11%)01 (11%)
J57 (41%)2 (22%)03 (33%)
λ15 (29%)001 (11%)
Untyped3 (17%)3 (33%)2 (66%)1 (11%)
  • a Monoclonal hybridomas were derived from unmanipulated 3H9.KI spleen cells from a single mouse 10 wk after cGVH induction. Ig-secreting hybridomas were assayed for dsDNA binding, persistence of the 3H9tg and L chain DNA rearrangement. Preliminary data from 3-wk hybridomas shows a similar skew to downstream L chain usage in the 3H9tg+ population. Fifty-three percent of these hybridomas are IgM and 47% are IgG. Both groups show a similar tendency in secondary L chain rearrangement in anti-dsDNA B cells.

  • b dsDNA binding was determined by a fluid-phase ELISA and confirmed by a solid-phase ELISA or Crithidia (relative values were consistent across all assays). The left portion of the chart (anti-dsDNA+) indicates dsDNA binding that gave an A405 ≥ 0.1 with Ab concentration (1.5 μg/ml). The right portion (anti-dsDNA) indicates no detectable binding.

  • c Retention of the 3H9 H chain tg was determined by PCR and corroborated by sequencing analysis. Positive clones are grouped in the column under 3H9tg+. Negative clones, with disrupted transgenes, are grouped in the column under 3H9tg.

  • d Anti-dsDNA+3H9+ includes two clones with double L chain rearrangement. Anti-dsDNA3H9 includes one clone with double rearrangement.

  • e Hybridomas were analyzed for L chain rearrangement. Genomic DNA was typed using the Schlissel degenerate Vκ forward primer (Vs) and the Jκ5 and Jκ2 reverse primer.