Splenic Immunohistochemistry of Clodronate Treated Mice

B6 recipients were treated or not with clodronate, followed by splenic harvest 24 hours later. Spleens were frozen, sectioned, and fixed as previously described.^{12 (link),22 (link)} Sections were then stained foe IgD, MARCO, and SIGNR1, then mounted using Prolong Gold antifade mounting media (ThermoFisher Scientific). A Leica SP8 multiphoton confocal microscope was used to capture images, with image analysis using Leica Application Suite software.

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Arthur C.M., Patel S., Sharma A., Zerra P.E., Chonat S., Jajosky R.P., Fasano R.M., Patel R., Bennett A., Zhou X., Luckey C.J., Hudson K.E., Eisenbarth S.C., Josephson C.D., Roback J.D., Hendrickson J.E, & Stowell S.R. (2022). Clodronate Inhibits Alloimmunization Against Distinct Red Blood Cell Alloantigens in Mice. Transfusion, 62(5), 948-953.

Publication 2022

Prolong Gold antifade mounting media SP8 multiphoton confocal microscope Application Suite software

Clodronate Confocal microscope Frozen Gold

Corresponding Organization : Harvard University

Other organizations : University of Virginia, Columbia University Irving Medical Center, Yale University

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Variable analysis

independent variables

Treatment with clodronate

dependent variables

Splenic macrophage marker expression (IgD, MARCO, and SIGNR1)

control variables

Tissue harvesting time (24 hours after treatment)
Tissue processing (freezing, sectioning, and fixing)
Immunostaining (IgD, MARCO, and SIGNR1)
Mounting media (Prolong Gold antifade)
Imaging (Leica SP8 multiphoton confocal microscope)
Image analysis (Leica Application Suite software)

controls

Negative control: B6 recipients not treated with clodronate

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