Journal:

Article Title: Nonselenium Glutathione Peroxidase in Human Brain

doi:

Figure Lengend Snippet: A: Electronmicrograph of a LB purified using magnetic beads and immunostained with a NSGP primary antibody and visualized with a secondary antibody conjugated to 12-nm gold. Top right inset shows the intact LB bound to magnetic beads (MB), small arrows indicate the boundary of LB. The area indicated was taken at higher power and shows the distribution of the gold particles. B: Immunoprecipitation of NSGP by anti-α-synuclein antibodies from a cytosol fraction of normal control human brain. Immunoblotting was conducted by using chicken IgY raised against recombinant NSGP, followed by biotinylated donkey anti-chicken IgY, and streptavidin-biotin-HRP complex. The membrane was developed using X-ray film and a enhanced chemiluminescence kit (Amersham Pharmacia Biotec, Sydney, Australia). Lane 1, Recombinant NSGP (∼15 ng). Lane 2, Precipitates using antibody to α-synuclein C-terminal 116 to 131 amino acids. Lane 3, Precipitates using antibody to α-synuclein N-terminal 11 to 26 amino acids. Lane 4, Precipitates from acetone-extracted cytosol using antibody to α-synuclein N-terminal 11 to 26 amino acids.

Article Snippet: Ultrathin sections were incubated for 3 hours with rabbit anti-NSGP and visualized with donkey anti-rabbit-conjugated 12-nm gold (Jackson ImmunoResearch).

Techniques: Purification, Magnetic Beads, Immunoprecipitation, Western Blot, Recombinant

Journal:

Article Title: Nonselenium Glutathione Peroxidase in Human Brain

doi:

Figure Lengend Snippet: A: PAGE analysis of soluble brain proteins from white and gray matter of normal, PD, and DLB brains. Lane 1, PD, FC, and GM; lane 2, PD, FC, and WM; lane 3, PD and SN; lane 4, NSGP recombinant protein; lane 5, DLB, FC, and GM; lane 6, DLB, FC, and WM; lane 7, N, FC, and GM; lane 8, N, FC, and WM; and lane 9, N and SN. Gel was stained with Coomassie blue. Molecular weights in kd are indicated at the left. Abbreviations: FC, frontal cortex; GM, gray matter; WM, white matter; SN, substantia nigra; DLB, dementia with LBs, N, normal. B: Western blot of the same samples as indicated in A. Primary antibody was protein A-purified rabbit anti-NSGP IgG, secondary antibody was a biotin-labeled donkey anti-rabbit (711-065-152, Jackson ImmunoResearch) and visualized with a Vector ABC kit using DAB.

Article Snippet: Ultrathin sections were incubated for 3 hours with rabbit anti-NSGP and visualized with donkey anti-rabbit-conjugated 12-nm gold (Jackson ImmunoResearch).

Techniques: Recombinant, Staining, Western Blot, Purification, Labeling, Plasmid Preparation

Journal:

Article Title: Nonselenium Glutathione Peroxidase in Human Brain

doi:

Figure Lengend Snippet: A–D: Low-power images (original magnifications, ×400) of the NSGP immunocytochemistry of gray and white gray matter from normal (A and B) and DLB (C and D) brain tissue. NSGP was visualized using a biotinylated donkey anti-rabbit secondary antibody and a Vector ABC staining kit with DAB. Sections were lightly counterstained with hematoxylin. NSGP-positive glial cells were markedly increased in both gray and white matter of DLB compared to normal. The arrows in C and D indicate NSGP-positive glial cells. E–G: Comparison of astrocyte labeling (oil immersion; original magnifications, ×1000) with NSGP antibodies in normal white cortical matter (E), and white (F) and gray matter (G) of DLB cortical tissue. H and I: LB labeling with rabbit anti-NSGP antibodies (H), and labeling with sheep anti-α-synuclein antibodies (I) within the gray matter of DLB tissue (oil immersion; original magnifications, ×1000). Immunoreactivity was visualized with biotinylated secondary antibodies and a Vector ABC kit using DAB. Arrows indicate the positively stained LBs within the cytoplasm of neurons. J–R: J to L show α-synuclein labeling with fluorescein isothiocyanate in homogeneous LBs (J) and two forms of concentric LBs (K and L); M to O show NSGP labeling with Cy5 in the same three LBs; and P to R depict the merged image showing the co-localization.

Article Snippet: Ultrathin sections were incubated for 3 hours with rabbit anti-NSGP and visualized with donkey anti-rabbit-conjugated 12-nm gold (Jackson ImmunoResearch).

Techniques: Immunocytochemistry, Plasmid Preparation, Staining, Labeling