High resolution fluorescence microscopy evidence on the transport of immunoglobulins. Differences between mammalian IgG, F(ab')2 and avian IgY

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Abstract

We describe the subcellular localization of horse F(ab')2 and IgG, and ostrich IgY labeled with fluorescein isothiocyanate (FITC) administered IV to mice. We used wide field high sensitivity fluorescence microscopy deblurred by 3-dimensional blind deconvolution of kidney, liver, lungs and brain sections. Sections were obtained from mice sacrificed 15 min, 1 or 5 h after receiving FITC-immunoproteins, counter-stained with DAPI (4′,6′-diamidino-2-phenylindole) and Evans blue. FITC-IgG and its fractions are rapidly taken up and extravasated by vascular endothelium. FITC-IgG and FITC-F(ab')2 appear to be quickly secreted by glomeruli endothelium and to be reabsorbed along all nephron segments. FITC-IgG and FITC-F(ab')2 appeared 15 min after IV injection within bronchial, alveolar and bile duct epithelium. Hepatocytes were loaded with fluorescence after 15 min of administration. Fluorescence was absent from brain slices, except for the endothelium of some vessels in brain ventricles which appeared intensely fluorescent. Fluorescence appeared in intracellular vesicles which conferred the tissues a glowing foamy aspect for up to 5 h after inoculation. Arterial elastic layers were intensely green after horse FITC-Ig inoculation. Ostrich FITC-IgY behaved completely differently to horse Ig's; only 1 h after injection it was possible to observe small brightly green scarce vesicles in vascular endothelium of arteries, interstitial kidney capillaries between nephron tubules and were also scarce in glomeruli endothelium; FITC-IgY appeared only in hepatic sinusoids in the liver. No IgY was seen in bronchial and alveolar endothelium, in bile ducts or in hepatocytes.

Highlights

We studied the subcelular distribution of IgY, F(ab')2 and IgG injected I.V. in mice. The Igs were labeled with fluorescein isotiocyanate and observed with deconvolution fluorescence microscopy. IgG and its F(ab')2 have a sub-cellular distribution very different from IgY. The Igs seem to pass kidney glomeruli but only IgG and F(ab')2 seem completely reabsorbed in nephrons. Our result that pharmacokinetics for IgG and F(ab')2 are very different of IgY PK.

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