Immunofluorescence of mouse inguinal lymph nodes

MH Michael Hamm
PS Pierre Sohier
VP Valérie Petit
JR Jérémy H. Raymond
VD Véronique Delmas
MC Madeleine Le Coz
FG Franck Gesbert
CK Colin Kenny
ZA Zackie Aktary
MP Marie Pouteaux
FR Florian Rambow
AS Alain Sarasin
NC Nisamanee Charoenchon
AB Alfonso Bellacosa
LS Luis Sanchez-del-Campo
LM Laura Mosteo
ML Martin Lauss
DM Dies Meijer
ES Eirikur Steingrimsson
GJ Göran B. Jönsson
RC Robert A. Cornell
ID Irwin Davidson
CG Colin R. Goding
LL Lionel Larue
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OCT-embedded lymph nodes were sectioned into 7-μm-thick transverse sections and rinsed in TBST, depigmented with H2O2 (Sigma, H1009) for 15 min, boiled for 20 min in 10 mM sodium citrate (VWR, 1120051000) and blocked with TBST containing 3% BSA (Sigma A9418). Sections were incubated overnight at 4 °C in TBST containing 3% BSA with antibodies against SOX10 (Abcam, ab155279). Sections were then incubated in secondary donkey anti-mouse 647 (Abcam, ab150107) and goat anti-rabbit IgG (H&L)-Affinity Pure, DyLight® 650 Conjugate (GtxRb-003-E650NHSX) antibodies for 2 h at RT. All sections were counterstained with DAPI (0.5 μg/mL in ethanol, Sigma, D9542). Images were captured using a ZEISS Axio Imager 2 with Axiocam 506 color cameras. Image analysis was performed using ZEISS ZEN, Adobe Photoshop, and ImageJ software.

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