Table 1.

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

FFE Fraction		Fraction of total vesicles that contain class II molecules		Fraction of class II–positive vesicles* that also contain BCR-internalized antigen		Fraction 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.