Immunohistochemistry, Image Analysis and Laser Scanning Confocal Microscopy was performed as previously described [50 (link)].
Analysis of CDK5 and pTau and pCRMP2 and pDCX was performed as previously described [119 (link)] by double labeling. For this purpose, coverslips or brain sections were incubated with a rabbit polyclonal primary antibody against p-Tau or p-CRMP2 detected with FITC-conjugated secondary antibodies [1:75, Vector Laboratories] and CDK5 or pDCX detected with Tyramide Red. All sections were processed under the same standardized conditions. The immunolabeled blind-coded sections were serially imaged with a laser scanning confocal microscope [MRC-1024; Bio-Rad] and analyzed with Image J v1.43 software [NIH, Bethesda, MD], as previously described [162 (link)]. For each case, a total of three sections were analyzed and four fields were examined. Results were expressed as percent of cells displaying CDK5 in the nucleus or levels of pixel intensity.
Analysis of CDK5 and pTau and pCRMP2 and pDCX was performed as previously described [119 (link)] by double labeling. For this purpose, coverslips or brain sections were incubated with a rabbit polyclonal primary antibody against p-Tau or p-CRMP2 detected with FITC-conjugated secondary antibodies [1:75, Vector Laboratories] and CDK5 or pDCX detected with Tyramide Red. All sections were processed under the same standardized conditions. The immunolabeled blind-coded sections were serially imaged with a laser scanning confocal microscope [MRC-1024; Bio-Rad] and analyzed with Image J v1.43 software [NIH, Bethesda, MD], as previously described [162 (link)]. For each case, a total of three sections were analyzed and four fields were examined. Results were expressed as percent of cells displaying CDK5 in the nucleus or levels of pixel intensity.
