The primary antibodies used to detect leukocytes in muscle cross sections were rat monoclonal anti-mouse 7/4 (1:20; AbD Serotec, Raleigh, NC) and rat monoclonal anti-mouse Ly-6C/G (clone RB6-8C5, 1:20; Invitrogen, Camarillo, CA).
Immunohistochemistry was performed as described previously (Schneider et al. 2012 (link)) except for the substrate reaction step. We applied Vector® VIP substrate solution (Vector® VIP Substrate Kit; Vector Laboratories, Burlingame, CA) to all sections for 3 min. Although the entire plantar flexor muscle group was harvested, the injury was evident primarily in the lateral gastrocnemius; therefore, only this area was used to compare E+ and E- groups quantitatively.
Immunohistochemistry was performed as described previously (Schneider et al. 2012 (link)) except for the substrate reaction step. We applied Vector® VIP substrate solution (Vector® VIP Substrate Kit; Vector Laboratories, Burlingame, CA) to all sections for 3 min. Although the entire plantar flexor muscle group was harvested, the injury was evident primarily in the lateral gastrocnemius; therefore, only this area was used to compare E+ and E- groups quantitatively.
