Journal: bioRxiv

Article Title: Optimal AAV capsid/promoter combinations to target specific cell types in the common marmoset cerebral cortex

doi: 10.1101/2024.05.09.593444

Figure Lengend Snippet: ( A ) Scheme to identify EGFP-expressing cell types by fIHC. Neurons and astrocytes were immunolabeled for NeuN and GFAP, respectively. Cells immunostained for Olig2 or Iba1 were identified as oligodendrocytes and microglia. Cells were detected with Hoechest 33342, NucBlue, in the mounting reagent ProLong Glass. ( B ) Representative immunohistochemical images of marmoset cerebral cortex that received injection of AAV2. Two serial slices were presented: one immunolabeled for GFP, NeuN, and GFAP, and another one for GFP, Olig2, and Iba1, as indicated at each panel. Scale bar, 100 μm.

Article Snippet: Tissue sections were reacted overnight at room temperature by immersion in following primary antibodies in blocking solution (2% Donkey Serum (S30-100ML, Merck), BSA (01862-87, Nacalai Tesque, Kyoto, Japan), 0.5% Triton X-100, 0.03% NaN3 in 1 x PB): rat monoclonal anti-GFP antibody (1:1,000; 04404-84; Nacalai Tesque, Kyoto, Japan), mouse monoclonal anti-NeuN antibody (1:1,000; MAB377; Merck), rabbit polyclonal anti-GFAP antibody (1:200; GFAP-Rb-Af800; Nittobo Medical, Tokyo, Japan), rabbit polyclonal anti-S-100β antibody (1:200; S100b-Rb-Af1000, Nittobo Medical), mouse monoclonal anti-Olig2 antibody (1:500; MABN50; Merck) and rabbit polyclonal anti-Iba1 antibody (1:500; 019-19741; Fujifilm Wako Chemicals, Tokyo, Japan).

Techniques: Expressing, Immunolabeling, Immunohistochemical staining, Injection

Journal: bioRxiv

Article Title: Optimal AAV capsid/promoter combinations to target specific cell types in the common marmoset cerebral cortex

doi: 10.1101/2024.05.09.593444

Figure Lengend Snippet: ( A ) Schema depicting the AAV genome structure. ( B - C ) Immunolabeled fluorescent images of EGFP in the cerebral cortex that received injection of AAV5 ( B ) or AAVrh10 ( C ) vectors expressing EGFP by the mMBP promoter. The middle immunofluorescence images present an overlay of immunolabeling for EGFP and the oligodendrocyte marker Olig2. The bottom images are magnifications of the boxed areas in the center images. Scale bar, 100 μm. ( D - E ) Summary graphs showing the specificity ( D ) and efficiency ( E ) of oligodendrocyte transduction. The dotted line in the graph indicates a ratio of oligodendrocytes to total cells present in the marmoset cortex (Figure S3). Error bars indicate S.E.M., and dots in the graph indicate the respective values for each of the individual marmosets. Asterisks indicate statistically significant differences between the AAV5 and AAVrh10. ** p < 0.01, *** p < 0.001 by student’s t-test.

Article Snippet: Tissue sections were reacted overnight at room temperature by immersion in following primary antibodies in blocking solution (2% Donkey Serum (S30-100ML, Merck), BSA (01862-87, Nacalai Tesque, Kyoto, Japan), 0.5% Triton X-100, 0.03% NaN3 in 1 x PB): rat monoclonal anti-GFP antibody (1:1,000; 04404-84; Nacalai Tesque, Kyoto, Japan), mouse monoclonal anti-NeuN antibody (1:1,000; MAB377; Merck), rabbit polyclonal anti-GFAP antibody (1:200; GFAP-Rb-Af800; Nittobo Medical, Tokyo, Japan), rabbit polyclonal anti-S-100β antibody (1:200; S100b-Rb-Af1000, Nittobo Medical), mouse monoclonal anti-Olig2 antibody (1:500; MABN50; Merck) and rabbit polyclonal anti-Iba1 antibody (1:500; 019-19741; Fujifilm Wako Chemicals, Tokyo, Japan).

Techniques: Immunolabeling, Injection, Expressing, Immunofluorescence, Marker, Transduction