Reflection contrast microscopy for high resolution detection of (3)H-estradiol in ultrathin sections of human stratum corneum.
A single autoradiographical method for light and electron microscopy (LM and EM) is presented. Human skin, containing (3)H-estradiol ((3)H-E2) after an in vitro permeation experiment, was processed via a non-extractive tissue preparation protocol, comprising cryo-fixation, freeze-drying, osmium tetroxide vapor fixation, and Spurr resin embedding. Semithin sections were processed for LM autoradiography, while ultrathin sections were processed both for high-resolution LM and EM autoradiography. The autoradiographs were visualized by bright-field microscopy (BFM), reflection contrast microscopy (RCM), and transmission electron microscopy to evaluate the potentials of RCM visualization in high-resolution LM autoradiography. RCM visualization of ultrathin vs. semithin resin sections showed an improved stratum corneum morphology. Histological staining was superfluous. The localization of (3)H-E2 in human stratum corneum using high-resolution LM autoradiography and RCM was as accurate as with high-resolution EM autoradiography.
Neelissen
J A
JA
AstraZeneca R&D Södertälje, Pharmacokinetcs and Biopharmaceutics, 151 85 Södertälje, Sweden. jan.neelissen@astrazeneak.com
Schrijvers
A H
AH
Junginger
H E
HE
Boddé
H E
HE
D001345:Autoradiography / Q000379:methods*; D015925:Cryopreservation; D004817:Epidermis / Q000737:chemistry* / Q000648:ultrastructure; D004958:Estradiol / Q000032:analysis*; D005612:Freeze Drying; D006801:Humans; D008854:Microscopy, Electron / Q000379:methods*; D010539:Permeability
D004958:Estradiol