Table 1 Sequence analysis of V gene rearrangements amplified from DLBCL and HRS cells of a composite lymphoma

Cells	Cells positive in PCR	PCR products amplified repeatedly	Original rearrangement potentially functional	Mutation frequency (%)	Functionality after somatic hypermutation	Ongoing mutation
DLBCL
CD20⁺ (2–4 cells/tube)	V_HL: 14/20	14 × V_H3–23^a	+	27–28^b	−	No
	V_HFRI^c: 0/20
	V_κ: 10/20	10 × V_κ1D–39	+	18	+	No
	V_λ: 1/20^d
	D_H: 7/10	7 × unrearranged
	KDE: 18/20	18 × unrearranged^e
Negative controls^f	V_HL, KDE, D_H: 0/4
	V_HL, KDE: 0/4
	V_HFRI, V_κ, V_λ: 0/8
CD20⁺ (1 cell/tube)	V_HL: 18/42	18 × V_H3–23	+	27–28^b	−	No
Negative controls^f	V_HL: 0/18
HL
CD30⁺ (1 cell/tube)	V_HL: 8/16	8 × V_H3–23	+	7	+	No
	V_HFRI: 14/20	14 × V_H3–23	+	7	+	No
	V_κ: 15/20^g	6 × V_κ1D–39^h	+	7	+	No
		14 × V_κ1D–6	+	8	+	No
	V_λ: 1/20^d
	D_H: 4/10	4 × unrearranged
	KDE: 13/16	10 × unrearranged
		11 × J_κ−C_κ intron–RSS/KDE rearrangement
Negative controls^f	V_HL, KDE, D_H: 0/4
	V_HL, KDE: 0/2
	V_HFRI, V_κ, V_λ: 2/8ⁱ

All sequences were deposited in the EMBL database under accession nos. AJ586890–896.
^aFrom two tubes in addition to the clonal rearrangement, unrelated unique rearrangements (V_H2–26 and V_H4–34) were amplified, likely due to the occasional micromanipulation of nontumor B cells from the DLBCL region.
^bIncluding one 1-bp insertion and three deletions (two 1-bp deletions and one larger deletion of 56 bp).
^cV_HFRI PCR was negative in DLBCL cells due to a large deletion encompassing the primer binding site in FRI.
^dOne V_λ1 PCR product out of 20 HRS cells and one V_λ3 PCR product out of 20 DLBCL cells were amplified but not sequenced. These amplificates likely represent cellular contamination, or picking of a bystander B cell in case of the analysis of CD20⁺ B cells.
^eOne single J_κ–C_κ intron–RSS/KDE rearrangement was amplified from a tube from which also a unique V_H rearrangement was amplified (see footnote a) and is thus likely derived from a nontumor B cell.
^fFor each 10 cells four aliquots of buffer covering the sections during micromanipulation were used as negative controls.
^gFrom two tubes in addition to the clonal rearrangement, unrelated unique rearrangements (V_κ1D–33 and V_κ1D–39) were amplified.
^hFive of six cells have both one V_κ1D–39 and one V_κ1D–6 rearrangement.
ⁱV_κ1D–33 and V_κ1–8 rearrangements not related to the tumor clone.

Search