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…combined with 20 µl of heat-inactivated human serum (inactivated at 56°C for 30 minutes) in 96-well U-bottom plates and incubated shaking at 37°C for 1 h. Subsequently, 10 µl of baby rabbit complement (Pel-Freez Biologics) was added to a final concentration of 10%, and plates were incubated shaking for 1 h at 37°C. All samples were tested in triplicate. Controls included bacteria alone, bacteria with serum only,…

…Frailty Indicator self-report questionnaire35. Serological analysis Functional MenACWY-specific antibody titers were determined using the serum bactericidal assay with baby rabbit complement (rSBA) (Pelfreez, lot: 21044-C) as described previously15,36. Target strains included MenA (strain 3125), MenC (strain C11), MenW (strain MP01240070), and MenY (strain S-1975). rSBA titers were expressed as the reciprocal…

…strains of pneumococcal serotype 19F, 18C, and 9V. Plates were incubated for 30 minutes on a shaker (750 RPM) at room temperature, followed by the addition of 10 μL baby rabbit complement (Pel-freeze biologicals) and differentiated HL-60 cells (cells: bacteria was 200:1). Plates were incubated for another 45 minutes at 37°C in a humidified chamber, followed by cooling on ice for 15 minutes to stop…

…adjusted to 10 5 CFU/mL. Following incubation at room temperature for 30 min with shaking, 40 µL of a differentiated HL-60 cell suspension containing 4 × 10 5 cells and 10 µL baby rabbit complement (BRC; Pel-Freez Biological: Rogers AR) was added. The cell mixture was incubated for 45 min at 37 °C with shaking. Next, 10 µL of the final mixture was spotted onto THY agar containing 1.5% Bacto agar (BD Biosciences).…

…to the manufacturer’s protocol. Neutrophils were adjusted to a concentration of 5 × 10 6 cells/mL in RPMI 1640 tissue culture media, and the bacteria were pre-incubated with baby rabbit complement (PelFreez, #31061) and heat-inactivated fetal bovine serum (FBS, Cat# 97068-085, VWR International). GBS strains were grown to the mid-logarithmic growth phase of OD 600 = ~0.4 and washed twice with sterile…

…assay The neutralizing activity of the mAbs and sera was determined using the MV or EV forms of the VACV Tian tan or WR strain. Neutralization of VACV EV was performed using a 10% baby rabbit complement (Pelfreez). For EV neutralization, MV was depleted with blocking mAbs at 20 mg/mL. BHK-21 cells were maintained in DMEM sup- plemented with 10% calf serum at 37°C, and 5% CO2. On the day of the experiment, BHK-21…