Improved immunolabelling of ultrathin cryosections using antibody conjugated with 1nm gold particles

During the last 5 years the diameter of the gold probes used for immuno-cytochemical staining at the electron microscopical (EM) level has been decreased. The advantage of small diameter gold probes is an overall increased labelling density. The disadvantage is a lower detectability due to the low electron density of smaller gold particles consequently an inconvenient high primary magnification needed for EM examination. Since 1 nm gold particles are barely visible by conventional EM examination the need for enlargement by silverenhancement of the gold particles has increased.In the present study of ultrathin cryosectioned material the results of immunostaining using 5 nm gold conjugated antibody and 1 nm gold conjugated antibodies are compared after silverenhancement of the 1 nm gold particles.Slices of freshly isolated mouse pituitary gland were immersion fixed for 20 min in 2 % glutaraldehyde /2 % paraformaldehyde. Blocks cryoprotected with 2.3 M sucrose were frozen in liquid nitrogen and ultra-cryosectioned on a RMC cryoultra-microtome.