Combination of fixation using PLP fixative and embedding in paraffin by the AMeX method is useful for histochemical studies in assessment of immunotoxicity.
To establish a method for processing lymphoid organs suited to morphological, immunohistochemical and enzyme histochemical analyses for assessment of immunotoxicity, we examined a combination of fixation with periodate-lysine-paraformaldehyde (PLP) fixative and embedding in paraffin by the AMeX method (PLP-AMeX method). Spleen and thymus removed from monkeys and rats were fixed in PLP fixative for 6 hours at 4 degrees C. After fixation, specimens were processed and embedded in paraffin by the AMeX method. In hematoxylin and eosin-stained sections, tissue architecture was well preserved. In immunohistochemical staining, markers of T lymphocytes (CD3, CD4, CD8), B lymphocytes (monkey: CD20cy, rat: CD45RA) and macrophage (monkey; CD68, rat: ED-1) were well identified according to their specificities, although the staining intensity of CD8 in the monkey and CD4 in the rat were somewhat weaker in PLP-AMeX-prepared sections than in those frozen. In enzyme histochemical staining, alkaline phosphatase activity was well preserved in neutrophils. In toluidine blue- and Giemsa-stained sections, eosinophil granules and the metachromasia of granules in basophil/mast cells were clearly detectable. These findings suggest that the PLP-AMeX method is a powerful tool for assessment of immunotoxicity.
Suzuki
Masami
M
Toxicology Laboratory, Chugai Pharmaceutical Co., Ltd., 1-135 Komakado, Gotemba-shi, Shizuoka 412-8513, Japan.
Katsuyama
Kiyoka
K
Adachi
Kenji
K
Ogawa
Yumie
Y
Yorozu
Keigo
K
Fujii
Etsuko
E
Misawa
Yasuyuki
Y
Sugimoto
Tetsuro
T
D000818:Animals; D005557:Formaldehyde; D007107:Immune System / Q000187:drug effects* / Q000473:pathology; D007150:Immunohistochemistry; D008239:Lysine; D008252:Macaca fascicularis; D016612:Paraffin Embedding / Q000379:methods*; D010504:Periodic Acid; D051381:Rats; D017207:Rats, Sprague-Dawley; D013154:Spleen / Q000187:drug effects / Q000473:pathology; D013950:Thymus Gland / Q000187:drug effects / Q000473:pathology; D016707:Tissue Fixation / Q000379:methods*; D015262:Xenobiotics / Q000633:toxicity*
D015262:Xenobiotics; C046311:periodate-lysine-paraformaldehyde; D010504:Periodic Acid; D005557:Formaldehyde; D008239:Lysine