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Table 1

Quantitative ImmunoEM Analysis of FFE Fractions Isolated from Antigen-pulsed A20 μWT Cells

FFE FractionFraction of total vesicles that contain class II moleculesFraction of class II–positive vesicles* that also contain BCR-internalized antigenFraction of BCR-internalized antigen-containing vesicles that are class II positive*
  %  %  % 
CIIV  75  71  58 
E/L  72  21§  34 
FFE FractionFraction of total vesicles that contain class II moleculesFraction of class II–positive vesicles* that also contain BCR-internalized antigenFraction of BCR-internalized antigen-containing vesicles that are class II positive*
  %  %  % 
CIIV  75  71  58 
E/L  72  21§  34 

CIIV- and endosome/lysosome (E/L)–containing FFE fractions from PC–OVA pulsed A20μWT cells were analyzed by multiple label immunoEM for the presence of class II molecules and BCR-internalized antigen (i.e., PC–OVA). The percent of vesicles labeled for one or both markers determined by an unbiased sampling technique (26). For CIIV FFE fractions, a total of 531 vesicle profiles (from four independent experiments) were analyzed. Values are reported as percentages.  

*

 Class II–positive endocytic vesicles represent either CIIV (CIIV FFE fractions) or class II–positive endosomes (endosome/lysosome FFE fractions).  

 In a parallel set of experiments, the distribution of a ligand (i.e., HRP-labeled goat anti-murine IgG) internalized via the endogenous muBCR of A20μWT cells was determined in CIIV FFE fractions. Quantitative analysis of these samples (292 total vesicle profiles) revealed that 90% of the vesicles that contained muBCR-internalized ligand were class II–positive structures (i.e., CIIV) and 85% of CIIV contain muBCR-internalize ligand.  

§

 The nonantigen-containing, class II–positive vesicles in the E/L-containing FFE fractions most likely represent contaminating, class II–containing Golgi-derived vesicles.  

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