Wetting agents for biological electron microscopy. I. General considerations and negative staining.

Abstract

SUMMARYThe most frequently used method of enabling aqueous suspensions of small, particulate, biological specimens to spread on the hydrophobic support films required for electron microscopy, is to add a surface‐active compound to the suspensions. The requirements of an ideal wetting agent for use in this situation are outlined. The polypeptide bacitracin has been found to meet all these requirements, and it is superior to the commonly‐used bovine serum albumin (BSA) in several respects. Bacitracin molecules are much smaller than those of BSA, and unlike BSA molecules they are not visualized by negative staining. Thus bacitracin produces a very even background against which the fine details of a specimen can be more easily interpreted. It facilitates the penetration of negative stain into small crevices and it produces a smaller halo of negative stain around specimen particles. Its use for spreading electron dense specimens is also demonstrated. Bacitracin is compatible with at least five commonly‐used negative stains, namely ammonium molybdate, potassium phosphotungstate (KPT), potassium silicotungstate, sodium tungstate and uranyl acetate. It is also biologically acceptable for use with a wide range of specimens. There is no evidence of bacitracin causing damage to specimens examined representing macromolecules, animal, plant and bacterial viruses, bacteria and subcellular organelles.