Journal: PloS one

Article Title: Kinetic analysis of mouse brain proteome alterations following Chikungunya virus infection before and after appearance of clinical symptoms.

doi: 10.1371/journal.pone.0091397

Figure Lengend Snippet: Figure 5. Western blot validations of differentially regulated proteins identified by 2D-DIGE and/or iTRAQ analyses. (A) Protein samples from each group used for proteomic analysis were minimally labeled with cyanine-3 dye. At the top, a representative protein profile of three biological replicates from brain lysates of mock (M), early (E), late paralytic (LP) and late tetanus-like (LT), separated by 10% SDS-PAGE is shown. WB with fluorescence-based methods was used to detect an overlaid fluorescent scan of the general protein patterns (Cy3 dye; green) and the specific immunoreactive proteins (FITC or Cy5 dye; red). To better visualize protein detection signals observed with each specific antibody used, corresponding cropped WB images are presented in grey levels. (B) The graphs correspond to the mean 6 S.D. of protein quantity measured by densitometry of the antigenic bands. Densitometry analyses were performed using TotalLab Quant v12.2 software (Nonlinear Dynamics), and data were normalized to levels of global protein pattern intensity. The values indicated under each graph correspond to fold changes from paired comparisons. The significance of the differential protein expression are indicated *, p,0.05; **, p,0.01; ***, p,0.001. A.U., arbitrary units. ANXA2: annexin A2; ARRB1: b-arrestin; GABRA1: c-aminobutyric acid receptor subunit alpha-1; GRASP1: GRIP-associated protein; ITGAV: integrin aV; MYPT1: myosin phosphatase target subunit 1; N-Ras: N-Ras; RABEP1: rabaptin-5; SYNGR3: synaptogyrin-3. doi:10.1371/journal.pone.0091397.g005

Article Snippet: Blots were saturated with 5% nonfat dried milk in PBS containing 0.05% (v/v) Tween 20 (PBS-T-milk) for 1 h. Western blot (WB) analyses were carried out with rabbit mono- or polyclonal antibodies directed against b-arrestin (1:5000, ARRB1, no. 4674, Cell Signaling Technology, Danvers, MA), GRIP-associated protein (1:500, GRASP1, no. sc-135681, Santa Cruz Biotechnology, Inc., Santa Cruz, CA), annexin A2 (1:100, ANXA2, no. sc-9061, Santa Cruz), integrin aV (1:100, ITGAV, no. 10179, Santa Cruz), myosin phosphatase target subunit 1 (1:100, MYPT1, no. sc25618, Santa Cruz), rabaptin-5 (1:1000, RABEP1, no. sc-15351, Santa Cruz), N-Ras (1:500, N-Ras, no. sc-519, Santa Cruz), synaptogyrin-3 (1:1000, SYNGR3, no. sc-68936, Santa Cruz), or with a goat polyclonal antibody directed against c-aminobutyric acid receptor subunit alpha-1 (1:100, GABAARa1 or GABRA1, no. sc-31045, Santa Cruz), diluted in PBS-T-milk and incubated overnight at 4uC.

Techniques: Western Blot, Multiplex sample analysis, Labeling, SDS Page, Fluorescence, Software, Expressing

Journal: Frontiers in Immunology

Article Title: Simian Immunodeficiency Virus Infection Mediated Changes in Jejunum and Peripheral SARS-CoV-2 Receptor ACE2 and Associated Proteins or Genes in Rhesus Macaques

doi: 10.3389/fimmu.2022.835686

Figure Lengend Snippet: Dynamics of different renin angiotensin system proteins, lactate dehydrogenase, and cytokine production in plasma/serum. (A) Plasma concentration of ACE2 (ng/mL) at different time points of infection. The plasma ACE2 gradually decreased from acute to chronic infection with the lowest concentration at 90 dpi, then recovered during the late chronic stage of infection. (B) Plasma concentration of Angiotensin II (Ang II, pg/mL) at different time points. Though not significant, post infection showed a slight increase of Ang II compared to the pre infection time point, except at 112 and 145 dpi. (C) Plasma concentration of angiotensin II receptor 1 (AGTR1) (ng/mL) during infection. A gradual decrease of plasma AGTR1 was detected from 14 dpi onward. (D) Serum LDH activity (U/L) in infected RMs and uninfected controls were evaluated using a Beckman Coulter AU 480 analyzer. No significant changes in serum LDH activity were detected across the different time points. Plasma IL-6 (E) , IL-1β (F) , and TNF-α (G) concentrations (pg/mL) at pre and post infection time points were evaluated by U-plex biomarker NHP multiplex assay. No significant changes were detected at any time points for any of the proinflammatory cytokines tested. The error bars represent the mean ± SE for each time point (n=10). Each symbol represents individual macaque in each plot. Asterisks indicate statistical differences between time points, as calculated by Bonferroni for ACE2 and Ang II and Tukey-Kramer for AGTR1 (* p < 0.05; ** p < 0.01; *** p < 0.001 and **** p < 0.0001).

Article Snippet: Inflammatory cytokines (IL-1β, IL-6, and TNF-α) and MCP-1 chemokine (monocyte chemoattractant protein-1) in plasma were quantified using a U-plex biomarker NHP multiplex assay (Meso Scale Diagnostics, USA) following manufacturer instruction with minor modification.

Techniques: Clinical Proteomics, Concentration Assay, Infection, Activity Assay, Biomarker Discovery, Multiplex Assay

Journal: International Journal of Molecular Sciences

Article Title: Colonic Mucosal Immune Activation in Mice with Ovalbumin-Induced Allergic Airway Disease: Association between Allergic Airway Disease and Irritable Bowel Syndrome

doi: 10.3390/ijms23010181

Figure Lengend Snippet: The results of cytokine ELISA assay. T-helper 1 (Th1) (IL-1β, interferon [IFN]-γ, and tumor necrosis factor [TNF]-α), Th2 (IL-4, IL-5 and IL-10), and proinflammatory cytokine (IL-6) levels were measured using ELISA with a multiplex protein assay kit. * p < 0.05, ** p = 0.064.

Article Snippet: A multiplex protein assay kit (Bio-Rad, Hercules, CA, USA) was used to measure the mucosal levels of Th1 (IFN-γ, IL-1β and TNF-α), Th2 (IL-4, IL-5 and IL-10), and proinflammatory cytokines (IL-6).

Techniques: Enzyme-linked Immunosorbent Assay, Multiplex Assay