Imaging Immune Response to GAS Infection

C57Bl/6 female mice (n = 3) were infected intra-dermally with 5×10⁷ M18 GAS and whole mount tissue sections and draining lymph nodes imaged 6 hours post infection. Whole mount tissue staining was carried out as described previously [2 (link)]. Frozen sections of lymph node were prepared by cryostat and fixed for 5 minutes in ice-cold acetone. Samples were stained with polyclonal anti-mouse LYVE-1 Ab and FITC-conjugated anti-group A carbohydrate antibody (Abcam). Whole mount tissue samples were viewed by Zeiss LSM780 confocal microscope, using either a Plan-Apochromat 10x/0.3DIC M27 (total magnification: 100x) or Plan-Apochromat 63x/1.4 oil (total magnification: 630x, resolution: 0.24 μm). For frozen lymph node sections, nuclei were counterstained with DAPI and sections were viewed on a Zeiss Axiovert S100 microscope equipped with epi-fluorescence.

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Lynskey N.N., Banerji S., Johnson L.A., Holder K.A., Reglinski M., Wing P.A., Rigby D., Jackson D.G, & Sriskandan S. (2015). Rapid Lymphatic Dissemination of Encapsulated Group A Streptococci via Lymphatic Vessel Endothelial Receptor-1 Interaction. PLoS Pathogens, 11(9), e1005137.

Publication 2015

LYVE-1 Ab Plan-Apochromat 63x/1.4 oil Axiovert S100 microscope

Acetone Antibody C57bl mice Carbohydrate Cold Confocal microscope Dapi Female Fitc Fluorescence Frozen sections Infection Lymph node Microscope Mouse Nuclei S100 Tissue

Corresponding Organization :

Other organizations : Imperial College London, MRC Human Immunology Unit, University of Oxford

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Variable analysis

independent variables

Infection route: intra-dermal
Infection dose: 5×10^7 M18 GAS

dependent variables

Tissue morphology and immune cell distribution in whole mount tissue sections
Immune cell distribution in draining lymph node sections

control variables

Mouse strain: C57Bl/6 female
Time point post-infection: 6 hours

controls

Positive control: Not mentioned
Negative control: Not mentioned

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