Immunofluorescence double labeling was performed as previously described.13 The primary antibodies (Ab) were goat anti‐Olig2 (1:200 dilution, AF2418, R&D Systems), rabbit anti‐Olig2 (1:200 dilution, ab109186, Abcam), rabbit anti‐Nogo‐A (1:100 dilution, ab62024, Abcam), and goat anti‐LCN2 (1:100, AF1757, R&D Systems). The secondary Abs were Alexa Fluor 488‐conjugated donkey anti‐rabbit mAb (1:500 dilution, Invitrogen), Alexa Fluor 488‐conjugated donkey anti‐goat mAb (1:500 dilution, Invitrogen), Alexa Fluor 594‐conjugated donkey anti‐rabbit mAb (1:500 dilution, Invitrogen), and Alexa Fluor 594‐conjugated donkey anti‐goat mAb (1:500 dilution, Invitrogen). Negative controls omitted the primary antibodies.
Olig2 is a helix–loop–helix transcription factor found in all OPCs, immature oligodendrocytes, and mature oligodendrocytes. Thus, Olig2+ cell numbers reflect all oligodendrocytes. In contrast, Nogo‐A identifies only the mature phenotype. Cells which were Nogo‐A−/Olig2+ cells were subclassified as oligodendrocyte precursor cells/immature oligodendrocytes.16 , 26 , 27
Olig2 is a helix–loop–helix transcription factor found in all OPCs, immature oligodendrocytes, and mature oligodendrocytes. Thus, Olig2+ cell numbers reflect all oligodendrocytes. In contrast, Nogo‐A identifies only the mature phenotype. Cells which were Nogo‐A−/Olig2+ cells were subclassified as oligodendrocyte precursor cells/immature oligodendrocytes.