Figure 5.

Agarose-acrylamide gel separation and Western blot analyses using mAb 3B3(−) and (+) of proteoglycan subpopulations extracted from the Hip (H) and Knee Joint cartilages (i.e. MK – medial; LK – lateral and MM – medial meniscus cartilages) of 3 month old (A) and 9 month old (B & C) retired breeder (RB) Hartley strain Guinea Pigs. Large aggregating proteoglycans (L – i.e. aggrecan subpopulations) and small leucine-rich proteoglycans (S) are identified by mAb 3B3 immunostaining either on the untreated nitrocellulose membranes [A & B where native 3B3(−) CS GAG chain epitopes are present – see Figure 2] or after the nitrocellulose membranes have been subjected to a chondroitinase ABC digestion [C] prior to their immunostaining with 3B3(+) where ‘CS-stubs’ containing a terminal unsaturated uronic acid adjacent to galactosamine-6-sulphate are present on both L and S proteoglycan subpopulations [C]. Note: only the slowest migrating (newly synthesized) aggrecan subpopulation is identified by immunostaining with 3B3(−) in [A] and [B]. The arrow points to the 3B3(−) positive staining that can only be found in the medial knee cartilage of 3-month-old animals.