HIGH-RESOLUTION PLATINUM-CARBON REPLICATION OF FREEZE-DRIED BASEMENT-MEMBRANE

High angle platinum/carbon (Pt/C) replication has proved to be a valua ble tool in analyzing basement membrane structure in human amnion, bov ine lens capsule, and the Engelbreth-Holm-Swarm (EHS) tumor. High reso lution replicas for transmission electron microscopy (TEM) have been a chieved by depositing 1.0 +/- 0.1 nm thick Pt/C films backed with rota ry deposited 12.5 +/- 2.5 nm thick carbon films. The basement membrane collagen IV network was observed to consist of fine branching filamen ts containing globular domains intrinsic to the filaments. A second qu asi-regular network is formed by laminin. Unidirectional 45 degrees an gle Pt/C replication was used for most of this work. The merits and de ficiencies of unidirectional vertical replication (80 degrees angle), unidirectional 45 degrees angle, and 20 degrees low angle rotary repli cation are discussed. Vertical replication produces the highest resolu tion replicas and has the potential for revealing the overall pattern of basement membrane structural assembly if basement membrane preparat ions freeze-dried in low salt can faithfully maintain their in vivo st ructure. (C) 1994 Wiley-Liss, Inc.