Journal: Brazilian Journal of Medical and Biological Research

Article Title: Recombinant 60-kDa heat shock protein from Paracoccidioides brasiliensis : is it a good antigen for serological diagnosis of paracoccidioidomycosis?

doi: 10.1590/1414-431X20175928

Figure Lengend Snippet: Western blot analysis of the recombinant proteins from P. brasiliensis recognized by sera from patients with paracoccidioidomycosis (PCM) or healthy subjects (NC). Recombinant proteins were separated by SDS-PAGE and transferred to nitrocellulose membranes. Membranes containing both proteins were incubated with sera from PCM patients ( A ) or NC subjects ( B ). After incubation with alkaline-phosphatase-conjugated antibody to human immunoglobulin, the reactions were revealed with BCIP-NBT. The arrowheads indicate the migration position of rPbHsp60 and rgp43. The identification of each patient serum is depicted by the numbers below each image. The negative reaction for both proteins is not shown. C , Relative number (%) of positive reactions with serum samples from PCM patients and NC group against rPbHsp60 and rgp43. No NC serum was reactive to rgp43.

Article Snippet: After the incubation, the membranes were washed five times and incubated with alkaline phosphatase-conjugated goat anti-human immunoglobulin antibody (isotyped M, G and A – whole molecule; Merck Millipore, USA), diluted 1:10,000 in a solution of 1% gelatin in TBS-T, for 1.5 hours at 37°C.

Techniques: Western Blot, Recombinant, SDS Page, Incubation, Migration

Journal: Frontiers in Immunology

Article Title: Complement Factor H Family Proteins Modulate Monocyte and Neutrophil Granulocyte Functions

doi: 10.3389/fimmu.2021.660852

Figure Lengend Snippet: FHR-1, FHR-5, FHL-1 and mini-FH bind to primary neutrophil granulocytes and monocytes. Binding of 300 nM of each protein (solid lines) to neutrophil granulocytes and monocytes in Ca 2+ and Mg 2+ containing DPBS was measured by flow cytometry. Filled histograms indicate autofluorescence, dotted lines show samples incubated without the proteins added (antibody controls). Binding of the added proteins (solid black line) was analyzed using polyclonal anti-FH Ab for FHR-1, FHL-1 and mini-FH detection, and monoclonal anti-FHR-5 for FHR-5 detection. A representative histogram out of three independent experiments is shown in each case.

Article Snippet: Bound proteins were detected via incubation with the polyclonal goat anti-human FH Ab in DPBS containing fetal bovine serum (FBS, 0.1%, EuroClone, Pero, Italy) for 15 minutes at 4°C, washed and labelled with Alexa-488-conjugated rabbit anti-goat antibody (Thermo Scientific, Waltham, MA) under the same conditions.

Techniques: Binding Assay, Flow Cytometry, Incubation