1. IgA and IgM VH repertoires in human colon: Evidence for clonally expanded B cells that are widely disseminated 
Wolfgang Holtmeier, Andreas Hennemann, Wolfgang F. Caspary 
Gastroenterology 
Volume 119, Issue 5, Pages (November 2000)
DOI: /gast
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2. Fig. 1
CDR3 length analysis of IgA VH transcripts in colon and peripheral blood of 4 healthy adults (A, B, C, and D). The bands indicated by numbers at the side of each gel were excised and directly sequenced without cloning. In addition, VH1, VH2, VH3, and VH4 bands from colon I and II (*) were excised, cloned, and than sequenced. Sequence analysis revealed multiple repetitive and clonally related transcripts at both sites (see Figures 2 and 7).
Gastroenterology  , DOI: ( /gast )
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3. Fig. 1
CDR3 length analysis of IgA VH transcripts in colon and peripheral blood of 4 healthy adults (A, B, C, and D). The bands indicated by numbers at the side of each gel were excised and directly sequenced without cloning. In addition, VH1, VH2, VH3, and VH4 bands from colon I and II (*) were excised, cloned, and than sequenced. Sequence analysis revealed multiple repetitive and clonally related transcripts at both sites (see Figures 2 and 7).
Gastroenterology  , DOI: ( /gast )
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4. Fig. 2
IgA VH sequences in colon and peripheral blood. Predominant bands indicated in Figure 1 from colonic and peripheral IgA VH repertoires were directly sequenced. Two x's on the right indicate identical CDR3 rearrangements in the 2 colonic biopsy sites or the 2 PBMC samples. Clonally related sequences were found in the 2 colon biopsy specimens (*) (see Figure 8). All sequences were in frame. IgA sequence data are available from EMBL/GenBank/DDBJ under accession numbers AF to AF
Gastroenterology  , DOI: ( /gast )
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5. Fig. 3
Clonality of IgA VH repertoires does not depend on B-cell numbers. VH6 transcripts from CD19+ and PBMC cell separations from 2 additional healthy subjects (E and F) are shown on the left. The CD19+ cell preparation contained approximately 10-fold more B cells than the PBMC separation (1X). Serial dilutions of cDNA from PBMC (40- and 400-fold) gave identical profiles. Colonic cDNA from subject A (right panel) was diluted up to 100-fold without significant change in the VH6 CDR3 profile.
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6. Fig. 4
CDR3 length analysis of IgM VH transcripts in colon and peripheral blood. The bands indicated by numbers at the side of each gel were excised and directly sequenced without cloning. Sequence analysis (see Figure 5) revealed identical or clonally related transcripts at both colon sites.
Gastroenterology  , DOI: ( /gast )
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7. Fig. 4
CDR3 length analysis of IgM VH transcripts in colon and peripheral blood. The bands indicated by numbers at the side of each gel were excised and directly sequenced without cloning. Sequence analysis (see Figure 5) revealed identical or clonally related transcripts at both colon sites.
Gastroenterology  , DOI: ( /gast )
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8. Fig. 5
IgM VH sequences in colon and peripheral blood. Predominant bands indicated in Figure 4 from colonic and peripheral IgM VH repertoires were directly sequenced. Two x's on the right indicate identical CDR3 rearrangements in the 2 colonic biopsy sites or the 2 PBMC samples. Clonally related sequences were found in the 2 colon biopsy specimens (*) (see Figure 8). All sequences were in frame. IgM sequence data are available from EMBL/GenBank/DDBJ under accession numbers AF to AF
Gastroenterology  , DOI: ( /gast )
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9. Fig. 6
Evidence for isotype-switched intestinal IgA B cells. (A) The CDR3 profiles of colonic IgM and IgA VH4–VH6 repertoires from subject C were compared on the same gel. Dominant bands of identical length that might contain IgM and IgA transcripts with identical CDR3 domains were excised and directly sequenced or cloned (IgA VH4 , asterisk). (B) The IgM and IgA VH6 transcripts tested were identical and, in the case of VH4, clonally related (see also Figure 7). Different CDR3 sequences were found when same length VH5 IgM and IgA transcripts were compared (IgM5021 and IgA5487).
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10. Fig. 7
Sequence analysis of cloned IgA VH transcripts obtained from single CDR3 bands. Eight individual CDR3 bands (marked by an asterisk in Figure 1) from 2 colon sites (I and II) were cloned and sequenced. Potential DH gene segments are underlined. Somatic mutations within the VH and JH regions are highlighted in gray. Nucleotide differences between clonally related sequences are boxed. Numbers on the right refer to the fraction of transcripts in each sample that carry the indicated junctional region. Identical or clonally related VH1, VH2, VH3, or VH4 transcripts were frequently present in both colonic sample sites. Sequence data are available from EMBL/GenBank/DDBJ under accession numbers AF to AF and AF to AF (VH2).
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11. Fig. 8
Protein sequences deduced from clonally related IgA or IgM transcripts recovered from distinct colonic sites. The amino acid sequences of the clonally related colonic IgA or IgM VH transcripts sequences given in Figures 2, 5, and 7 are shown. Amino acid replacements that were encoded from different nucleotides are boxed. Most of the nucleotide substitutions (n = 76) were silent (51%) or resulted in amino acid replacements with similar chemical properties (20%).
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12. Fig. 9
JH usage of intestinal and peripheral IgA transcripts. The data was calculated from the IgA sequences given in Figures 2 and 7.
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