Immunohistochemistry to Study the Relationship Between Macrophages and Infiltrated Immune Cells
Published: February 29, 2024
Abstract
Source: Albano-Aluquin, S. et al., An Immunohistopathologic Study to Profile the Folate Receptor Beta Macrophage and Vascular Immune Microenvironment in Giant Cell Arteritis. J. Vis. Exp. (2019)
This video demonstrates a technique to quantify the relative ratio of macrophages among the total infiltrated immune cell populations in temporal artery biopsy. The immunohistochemical staining of a tissue section affected by autoimmune disease selectively stains the immune cells with specific receptors to visually differentiate them from the rest of the cells.
Protocol
All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines. 1. Histopathological Preparation, and Processing After Obtaining Temporal Artery Biopsy NOTE: The temporal artery biopsy (TAB) is performed under local anesthesia and sterile conditions by a certified surgeon. After surgically obtaining a 3 cm arterial section on the more affected side, the specimen is fixed in fo…
Divulgations
The authors have nothing to disclose.
Materials
| Microtome | Reichert-Jung | ||
| Plain and frosted microscope slides and cover slips | Fisher Scientific | ||
| Vertical rack | Fisher Scientific | ||
| Light microscope | Olympus BX60 microscope | ||
| Xylene | |||
| Acetone in distilled water | concentrations 100%, 90%, 70% | ||
| Tris-buffered saline solution (TBS) | Dako Wash BufferS3006 | ||
| 3% hydrogen peroxide in TBS | |||
| Diaminobenzidine (DAB) | Sigma Fast Tablet set | ||
| Chemical Permount Mounting Medium | Fisher Scientific | SP-15-100 | |
| Harleco Gill's III Hematoxylin | Fiisher Scientific 23-750-019 | ||
| Harleco Eosin Y 1% Alcoholic Stock Solution | Fisher Scientific | 23-749-977 | |
| 10 mM citric acid monohydrate (pH 6.0) | |||
| Polyclonal rabbit anti-folate receptor beta | Manohar Ratnam | optimized at 1:800 | |
| Monoclonal mouse anti-human CD68 | Dako | M071801 | optimized at 1:200 |
| Polyclonal rabbit anti-CD3 | Agilent Dako | A0452 | optimized at 1:100 |
| Polymer goat anti- rabbit/mouse secondary antibody | Dako | ||
| Placental tissue | for FRB positive control |
Tags
Citer Cet Article
Immunohistochemistry to Study the Relationship Between Macrophages and Infiltrated Immune Cells. J. Vis. Exp. (Pending Publication), e21983, doi: (2024).