Journal: Biosensors

Article Title: Development and In-Depth Characterization of Bacteria Repellent and Bacteria Adhesive Antibody-Coated Surfaces Using Optical Waveguide Biosensing

doi: 10.3390/bios12020056

Figure Lengend Snippet: ( A ) Testing the binding abilities of different polyclonal Abs with an in-house ELISA test with different amounts of intact E. coli cells as antigen. Black: Bio-Rad 4329-4906, white: Thermo Fisher PA1-7213, grey: Thermo Fisher PA1-73032. ( B ) The binding ability of a monoclonal Ab (Bio-Rad OBT0749) with an in-house ELISA test with different amounts of intact E. coli cells as antigen. For detection, biotinylated polyclonal Ab (Bio-Rad 4329-4906) was used. The inset shows the schematic drawing of the assay.

Article Snippet: Based on the above results, we chose the polyclonal Bio-Rad rabbit 4329-4906 Ab deposited on protein A surfaces and PAcrAM-P blocking for further experiments with bacteria-specific detection.

Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay

Journal: Biosensors

Article Title: Development and In-Depth Characterization of Bacteria Repellent and Bacteria Adhesive Antibody-Coated Surfaces Using Optical Waveguide Biosensing

doi: 10.3390/bios12020056

Figure Lengend Snippet: The measured surface mass densities of the Ab layers at the maximum sensor signal and after washing off the reversibly bound protein mass. Various immobilization strategies were employed and are marked in the figure. P: polyclonal, M: monoclonal.

Article Snippet: Based on the above results, we chose the polyclonal Bio-Rad rabbit 4329-4906 Ab deposited on protein A surfaces and PAcrAM-P blocking for further experiments with bacteria-specific detection.

Techniques:

Journal: Biosensors

Article Title: Development and In-Depth Characterization of Bacteria Repellent and Bacteria Adhesive Antibody-Coated Surfaces Using Optical Waveguide Biosensing

doi: 10.3390/bios12020056

Figure Lengend Snippet: Typical OWLS kinetic curves of Ab deposition and their corresponding fits are shown. ( A ) Polyclonal Ab (Bio-Rad rabbit 4329-4906 on protein A surface). ( B ) Monoclonal Ab (Bio-Rad mouse OBT0749 on protein A surface). The smaller adsorbed surface mass density and slower adsorption are clearly visible in the case of the monoclonal Ab.

Article Snippet: Based on the above results, we chose the polyclonal Bio-Rad rabbit 4329-4906 Ab deposited on protein A surfaces and PAcrAM-P blocking for further experiments with bacteria-specific detection.

Techniques: Adsorption

Journal: Biosensors

Article Title: Development and In-Depth Characterization of Bacteria Repellent and Bacteria Adhesive Antibody-Coated Surfaces Using Optical Waveguide Biosensing

doi: 10.3390/bios12020056

Figure Lengend Snippet: Scatter plots of data resulted from the kinetic fits. ( A ) The monoclonal and polyclonal Abs can be easily distinguished based on the adsorption rate constant ( k a ). ( B ) The different protein A and MG surfaces are very distinct, and additionally, the groups of polyclonal and monoclonal Abs are separated well from each other based on the dissociation rate constant ( k d ). ( C ) The polyclonal and monoclonal Abs groups are separated based on the irreversible association rate constants ( k i ). ( D ) Additionally, polyclonal and monoclonal Abs separate into different groups when the footprints of reversibly and irreversibly adsorbed molecules are considered.

Article Snippet: Based on the above results, we chose the polyclonal Bio-Rad rabbit 4329-4906 Ab deposited on protein A surfaces and PAcrAM-P blocking for further experiments with bacteria-specific detection.

Techniques: Adsorption

Journal: Biosensors

Article Title: Development and In-Depth Characterization of Bacteria Repellent and Bacteria Adhesive Antibody-Coated Surfaces Using Optical Waveguide Biosensing

doi: 10.3390/bios12020056

Figure Lengend Snippet: Bacterial adsorption on the Ab-coated sensor surfaces and the detection limit of OWLS. ( A ) Sensogram of a complete experiment, including the in situ coating procedures and subsequent bacteria adsorption for protein A-based immobilization and PAcrAM-P blocking. ( B ) E. coli adsorption on polyclonal Ab-coated surfaces using PAcrAM-P blocking; a 10 9 cells/mL concentration was employed. The signals with statistics are clearly distinguishable from the relevant control signal (full layer without Ab). ( C ) OWLS signal for a series of bacterial concentrations using protein A-based immobilization with PAcrAM-P blocking.

Article Snippet: Based on the above results, we chose the polyclonal Bio-Rad rabbit 4329-4906 Ab deposited on protein A surfaces and PAcrAM-P blocking for further experiments with bacteria-specific detection.

Techniques: Adsorption, In Situ, Bacteria, Blocking Assay, Concentration Assay, Control

Journal: bioRxiv

Article Title: Evolutionary engineering a larger porin using a loop-to-hairpin mechanism

doi: 10.1101/2023.06.14.544993

Figure Lengend Snippet: (A, B) Western blots with the nitrocellulose membrane stained with Ponceau S and subsequently immunoblotted against various antibodies. (A) Western blot of different fractions of E. coli displaying the localization of the his-tagged recombinant proteins PorB WT and PorB 18 in the outer membrane. SurA, a periplasmic protein, indicates the localization of periplasmic proteins. Whole-cell fractions (WC), soluble (S), inner membrane and trapped periplasmic protein (IM/P), and outer membrane (OM) (B) Western blot displaying the susceptibility of proteins to proteinase K under different conditions. His-tagged PorB WT (blue asterisks) and PorB 18 (green asterisks) show resistance to proteinase K when not boiled but are partially susceptible when heat denatured. TolC (red asterisks) is used as a positive control as it shows a 5 kDa cleavage in the presence of proteinase K resulting in a 45 kDa cleavage product (red arrows) .

Article Snippet: The following antibodies were diluted as per manufacturer recommendation with TBST with 1% gelatin: THE Anti-His tag monoclonal mouse antibody (GenScript), anti- E. coli SurA polyclonal rabbit antibody (Cusabio), IRDye 800CW goat anti-mouse IgG secondary antibody (LI-COR Biosciences), IRDye 680RD donkey anti-rabbit IgG secondary antibody (LI-COR Biosciences). anti- E. coli TolC was purified from rabbit sera and diluted to 1:2500 with 1% gelatin in TBST.

Techniques: Western Blot, Membrane, Staining, Recombinant, Positive Control

Journal: EBioMedicine

Article Title: Gut bacterial type III secretion systems aggravate colitis in mice and serve as biomarkers of Crohn's disease.

doi: 10.1016/j.ebiom.2024.105296

Figure Lengend Snippet: Fig. 1: A. pulmonis mAT1 harbors a functional T3SS. (A) Phylogenetic tree of A. pulmonis mAT1 (red), compared to A. dolens LMG 26841 strain and 14 closest type strains automatically picked by the Type Strain Genome Server (TYGS). The phylogenetic tree was generated by TYGS with default settings. (B) 35 online-available Achromobacter genomes were re-annotated by NR and searched for genes of T3SS proteins that belong to each T3SS ortholog. The quantities of each T3SS ortholog were plotted in the heatmap. Achromobacter strains were classified into host- isolated and environmental-isolated groups. Achromobacter strains in each group were organized by phylogenic tree generated by GTDB-Tk and IQ-TREE2 software. Achromobacter strains that contain more than 10 T3SS orthologs annotated by NR are considered T3SS-harboring strains and labeled red. (C) Neighbor-joining phylogenetic tree T3SS ATPase SctN of representative T3SS-harboring bacteria strains. Bacteria names are color-coded to indicate designated T3SS families. (D) T3SS gene cluster of A. pulmonis mAT1 compared with that of B. bronchiseptica RB50. Genes are color-coded to indicate designated components of the T3SS apparatus. (E) The relative expression level of T3SS genes of A. pulmonis mAT1 grown in designated media, measured by qRT-PCR. All data are expressed as the mean ± SEM of at least three experimental repeats, each with two technical replicates. The Kruskal–Wallis test with Dunne’s multiple comparisons was performed and adjusted by the Benjamini-Hochberg method. For Dunn’s multiple comparisons test, the mean of each group was compared with the mean of the control group. *p < 0.05, **p < 0.01, ***p < 0.001. (F) Immunoblotting of secreted AopD by designated strains. Each bacterial culture was centrifuged

Article Snippet: The proteins in the supernatant fraction were precipitated in 10% (v/v) trichloroacetic acid.26 Proteins in both fractions were separated via 12% SDS-PAGE and analyzed by immunoblotting with a rabbit anti-AopD polyclonal antibody and a rabbit anti-DnaK polyclonal antibody (CUSABIO; #CSB-PA633459HA01EGW).

Techniques: Functional Assay, Generated, Isolation, Software, Labeling, Bacteria, Expressing, Quantitative RT-PCR, Control, Western Blot

Journal: eLife

Article Title: Colicin E1 opens its hinge to plug TolC

doi: 10.7554/eLife.73297

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-SurA Rabbit polyclonal (rabbit polyclonal) , Cusabio , Cat#CSB-PA359693HA01ENV; RRID: E0109A , (1:2500).

Techniques: Recombinant, Software