Immunology and Infection
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Phenotypic Characterization of Macrophages from Rat Kidney by Flow Cytometry
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Summary October 18th, 2016
This manuscript describes a detailed protocol for phenotypic and quantitative analysis of resident macrophages from rat kidneys by flow cytometry. The resulting stained cells can be also used for other applications, including cell sorting, gene expression analysis or functional studies, thus increasing the information obtained in the experimental model.
Transcript
The overall goal of this procedure is to quantify and characterize resident macrophages from rat kidneys by flow cytometry.This method can have answer key question in the fluidity such what is the contribution of each macraphage of type in kidney injury.The main advantage of this technique is that it allows the kidney catheterization of differing protein marker.Thus, limiting the of cellular integrity of Demonstrating the procedure will be Elena Olivares, A.P.A.S.D.student from the Complutense University and Melania Guerrero, A.P.A.S.D.student from For kidney perfusion and extraction, begin by pinching a small fold of skin to confirm that the rat has reached the appropriate level of sedation.And cover the eyes with Vet ointment.Place the animal on a surgical table in the supine position.And soak the abdomen with 70%ethanol.Next, make a central incision through the abdominal skin and peritoneum from the pubis to the ribcage to expose the pleural and abdominal cavities.Use the perfusion system to inject 0.9%saline solution into the abdominal aorta to flush the kidneys.Cutting the aorta at the abdomen to help release the blood.When all of the blood has been removed from the kidneys, sever the renal hilum.Then press the edge of each kidney to carefully decapsulate the organs.And transfer the extracted kidneys into fresh H.B.S.S.To generate a single kidney cell suspension, use scissors to cut half of one kidney into small fragments.Transfer the tissue pieces into a 1.5 mL tube and add 1 mL of freshly prepared collagenase to the tube for a 30 minute incubation at 37
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