Journal: bioRxiv

Article Title: The restricted nature of protein glycosylation in the mammalian brain

doi: 10.1101/2020.10.01.322537

Figure Lengend Snippet: Protein lysate from mouse cortex and cerebellum with human plasma as a positive control was treated with or without PNGase F and visualized using biotinylated lectins (Con A, GNL, PHA-E, AAL, RCA, and SNA) in addition to immunoblotting for actin and staining for total protein. Non-specific binding of lectins to PNGase F is noted by an asterisk (*) near 35 kDa. A schematic with common lectin-binding sites is shown for reference.

Article Snippet: Membranes were then incubated in 5% BSA in TBS-Tween 0.1% for 1 hour, followed by incubation with biotinylated lectins (Vector Labs: AAL B-1395, SNA B-1305, GNL B-1245, PHA-E B-1125, RCA B-1085, Con A B-1105) at a 1:1,000 dilution (1:20,000 for Con A) and 1:2,000 dilution of mouse anti-actin antibody (Abcam, ab8226) in 5% BSA in TBS-Tween 0.1%, overnight at 4°C on a rocking platform shaker.

Techniques: Positive Control, Western Blot, Staining, Binding Assay

Journal: Mucosal Immunology

Article Title: Secretory IgA N -glycans contribute to the protection against E. coli O55 infection of germ-free piglets

doi: 10.1038/s41385-020-00345-8

Figure Lengend Snippet: Pooled human colostrum/milk served as the source of SIgA. a Gel-permeation chromatography-purified SIgA was separated by SDS PAGE stained by Coomassie Blue. b SIgA digested with C. ramosum protease generates a large SIgA fragment designated Fc-SC (containing Fc portions of IgA plus SC plus J chain) and Fab fragment. Fc-SC and Fab were separated by gel-permeation chromatography and analyzed by SDS PAGE under non-reducing and reducing conditions to identify components of Fc-SC (SC and constant region—cα) and Fab (LC light chain and vα variable region). c SIgA deglycosylated by EndoH and PNGaseF was designated dg-SIgA and characterized by the mobility shift on SDS PAGE in comparison to untreated (N) SIgA (left part of c ) and by reduction of reactivity with high mannose-specific lectin from Galanthus nivalis on Western blot (right part of c ).

Article Snippet: The rate of N -glycans removal was determined on western blot with biotin-conjugated Galanthus nivalis lectin (Vector Laboratories, Burlingame, CA) and HRP-neutravidin as the chemiluminescence reduction recorded on cooled CCD camera.

Techniques: GPC Assay, Purification, SDS Page, Staining, Mobility Shift, Western Blot