Preparation of a Rabbit Anti-Guinea Pig Basophil Serum: in Vitro and in Vivo Characterization

Abstract

Abstract A high titer anti-basophil serum was prepared by immunizing rabbits with partially purified guinea pig peripheral blood basophils. Successive absorptions with normal guinea pig serum, red blood cells, polymorphonuclear leukocytes, and peritoneal-exudate macrophages and lymphocytes rendered the anti-basophil serum (ABS) reactive with basophils but no other leukocytes as determined by immunofluorescence, immunoferritin electron microscopy, and cytotoxicity assay. The basophil antigen(s) was expressed on the cell surface in close proximity to the plasma membrane and could be capped. ABS induced complement (C)-independent basophil degranulation and histamine release, but C was required for substantial basophil cytotoxicity and maximal degranulation and histamine release. ABS may recognize a basophil antigen(s) shared with tissue mast cells, because intradermal administration of small amounts of ABS produced local mast cell degranulation, reduction in skin histamine content, and an increase in vascular permeability that could be partially blocked by promethazine. ABS was tested in vivo in guinea pigs sensitized both with ovalbumin for cutaneous basophil hypersensitivity (CBH) and with mycobacteria for classic delayed hypersensitivity (DH). Administration of 0.25 to 1.0 ml of ABS i.v. reduced circulating basophils to 30% control values (p < 0.05) at 2 hr and reduced the number of bone marrow basophils to 7% control values (p < 0.001) at 24 hr. Other marrow and circulating leukocytes were not significantly affected. ABS also markedly reduced the number of basophils infiltrating 24-hr CBH reactions as determined by histology and histamine content. Other leukocytes were not altered nor was the cellular composition of DH or Con A-induced reactions affected. Systemic administration of ABS had no appreciable effect on subepidermal mast cells: the number and morphology of subepidermal mast cells, and the histamine content of normal skin were the same in animals that had received either i.v. ABS or NRS 24 hr before sacrifice. The CBH reactions in ABS-treated animals were more intensely erythematous than those in NRS-treated controls (p = 0.015). ABS may prove useful in analyzing the basophil's role in inflammatory reactions.