Immunoblots were prepared with extracts from 80% confluent cell cultures lysed in RIPA buffer (Pierce, Rockford, IL, USA) containing protease and phosphates inhibitors (Calbiochem). Cell lysates were centrifuged at 14,000 rpm for 15 min and resolved by SDS-PAGE. Immunoblots were prepared and visualized as previously described, using MAb 9.2 to detect CEACAM1-4S [13] . The density of the MAb 9.2 reactive bands was determined by image analysis of digital images of immunoblots captured using a Versadoc Imaging System (Bio-Rad) and Quantity One software (Bio-Rad), as previously described [13] .