Allophycocyanin (APC)-conjugated anti-rat primary antibodies are directly labeled immunoreagents that bind specific rat antigens and carry the APC fluorophore, enabling single-step detection in flow cytometry and related assays. They are distinct from APC-conjugated anti-rat secondary antibodies, which are raised in another host to detect rat immunoglobulins.
Introduction
APC is a phycobiliprotein with an excitation maximum of approximately 650 nm and an emission maximum of approximately 660 nm, making it optimally excited by the 633–640 nm red laser line standard on most modern flow cytometers. The large Stokes shift and far-red emission minimize spectral overlap with green (FITC) and orange (PE) fluorophores, facilitating multicolor panel design. Direct conjugation to a primary antibody eliminates the secondary antibody incubation step, reducing assay time and avoiding cross-reactivity from anti-rat secondary reagents—a particular advantage in rat-on-rat applications where endogenous immunoglobulins would otherwise produce high background.
Applications
APC-conjugated anti-rat primary antibodies are validated primarily for flow cytometry, with representative clones including anti-rat CD4 (clone W3/25, recognizing a 55 kDa glycoprotein on T helper cells and macrophages), anti-rat CD4 (clone OX35), anti-rat CD11b/c (clone OX-42, reacting with the CR3 complement receptor on monocytes, granulocytes, macrophages, dendritic cells, and NK cells), anti-rat CD71 (clone OX-26, targeting the transferrin receptor), and anti-rat CD44H (clone OX-49, recognizing the standard isoform of CD44). These conjugates are quality-tested by immunofluorescent staining with flow cytometric analysis, with a suggested use of ≤0.25 µg per million cells in 100 µL volume. Additional reported applications for some clones include immunohistochemistry on acetone-fixed frozen sections and functional blocking—for example, clone OX-42 has been shown to block complement-mediated rosette formation and leukocyte migration, and clone W3/25 inhibits IL-2 production by T helper cells and prevents autoimmune T cell transfer in an MBP-induced EAE model.