Transmission electron microscopical studies on some haemolymph proteins from the marine polychaete Nereis virens

The hexagonal bilayer haemoglobin molecule from Nereis virens has been inve stigated in a comparative study using several different negative stain elec tron microscopical specimen preparations (i.e. by conventional adsorption t o continuous carbon support films, by the negative staining-carbon film tec hnique and by negative staining across the holes of holey carbon support fi lms with air-drying and rapid freezing/cryo-negative staining). The benefit s and limitations of these different approaches are indicated, with the ove rall conclusion that negative staining with ammonium molybdate across holes creates the best possibilities for molecular imaging, and also has the pot ential for the creation of two-dimensional (2D) crystals/arrays at the flui d-air interface. Of the different negative staining procedures presented, c ryo-negative staining reveals the greatest details of N. virens haemoglobin . This is exemplified by the direct visualisation of the central linker-ass embly within the haemoglobin molecule, a structural feature less clearly de fined by the other negative staining techniques. A discoidal lipoprotein mo lecule (diameter 30-60 nm; thickness ca 8 nm) has been detected in N. viren s, which represents the first documented account of an annelid haemolymph l ipoprotein. The biological implications of this lipoprotein for lipid trans port remain to be established. The presence of a low concentration of ferri tin molecules in N. virens haemolymph is also shown, assisted by the format ion of small 2D ferritin arrays in negatively stained specimens prepared ac ross holes. (C) 2001 Elsevier Science Ltd. All rights reserved.