Canine T cells were collected from leukapheresis products obtained from peripheral blood of healthy research dogs at the University of Pennsylvania, Veterinary School of Medicine, with Institutional Animal Care and Use Committee (IACUC) approval. The cells were cultured and expanded with cell-based artificial antigen-presenting cells (aAPCs) as described before.39 (link) In brief, the human erythroleukemic cell line K562 transduced with lentiviral vector to stably express human FcγRII (CD32) and canine CD86 was used as artificial APCs, which were provided by Nicola Mason (School of Veterinary Medicine, University of Pennsylvania). Before expanding canine T cells, aAPCs were irradiated with 10,000 rad and washed with R10 media. Canine T cells were cultured with aAPCs at a 2:1 ratio to a final concentration of 1 × 106 canine T cells and 5 × 105 aAPCs per mL with 0.5 μg/mL mouse anti-canine CD3 (Bio-Rad) in R10 media with 30 IU/mL rhIL-2. The concentration of the expanding canine T cells was calculated on a Coulter Multisizer (Beckman Coulter) and maintained at 1.0–2.0 × 106 per mL R10 media with rhIL-2.
Coulter multisizer
Manufactured by Beckman Coulter
Sourced in United States
The Coulter Multisizer is a laboratory instrument designed for particle size analysis. It uses the Coulter Principle to measure the size and count of particles suspended in an electrolyte solution. The instrument provides precise and accurate particle size distribution data, which is essential for various applications in industries such as pharmaceuticals, materials science, and environmental monitoring.
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Lab products found in correlation
6 protocols using coulter multisizer
1
Expansion of Canine T Cells
2
T Cell Transduction and Culture Protocol
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Human T cell transduction and culture were performed as previously described.9 Briefly, isolated T cells were derived from leukapheresis products obtained from the Human Immunology Core at the University of Pennsylvania using de-identified healthy donors under an institutional review board-approved protocol. T cells were stimulated with Dynabeads Human T-Activator CD3/CD28 (Life Technologies) at a bead-to-cell ratio of 3:1. After 24-hr stimulation, lentivirus was added into the culture media and thoroughly mixed to produce stably transduced CAR T cells. The concentration of the expanding human T cells was calculated on a Coulter Multisizer (Beckman Coulter) and maintained at 1.0–2.0 × 106 per mL in R10 media supplemented with 30 IU/mL recombinant human IL-2 (rhIL-2; Proleukin, Chiron). Stably transduced human CAR T cells used in the in vivo study were normalized to 30% CAR+ before transplantation.
3
Karlotoxin Effects on Prorocentrum donghaiense
4
Characterization of Ultrasound-Exposed DPMC
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DPMC with US exposure was additionally characterized via dynamic light scattering (DLS) and transmission electron microscopy (TEM). Briefly, 1 mL DPMC in a 2.0 mL Eppendorf tube was placed in a water tank. Exposure was achieved with 20 mm US probe of a therapeutic US system (CZ906A, Chongqing Medical University, Chongqing, China), which was placed 2 cm away from the Eppendorf tube below the water surface. Irradiation parameters were set as follows: 1 MHz, 2 % duty cycle, duration of 1 min and US intensity of 1 W/cm211 (link), 12 (link). To characterize the morphologic differences between DPMC with and without US exposure, the suspension was mounted on a slide with a coverslip and visualized using confocal microscopy. The size distribution of DPMC was further estimated using a Coulter Multisizer (Beckman Coulter, Brea, California, USA). To determine the drug loading content of DPMC, the amount of DP that bound to MBs was estimated by removing unbound DP via centrifugation at 400 g for 3 min 9 (link). Unloaded DOX in the collected solution was quantified at 480 nm with a UV-Vis spectrophotometer, and the drug loading content calculated using the formula: DLC (%) = (total added drug - unloaded drug)/total amount of MB.
5
Biofilm-Forming S. aureus Culturing
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S. aureus Phillips, a biofilm forming bacteria, was used in this study. The strain was isolated from a patient diagnosed with osteomyelitis and has been extensively studied [9 (link),10 (link),33 (link)]. S. aureus cultures were started by inoculation (10 μL) from glycerol stocks into tryptic soy broth (TSB; 50 μL) supplemented with 0.25% (w/v) glucose and grown at 37°C with constant rotation at 141 rpm in shake flasks. The growth of the bacterial strains was monitored by measuring the absorbance of the broth at 600 nm on a spectrophotometer. The cells were harvested at mid-exponential phase (OD600 = 0.3 to 0.35), centrifuged (4000 rpm) for 15 min at 4°C and resuspended in phosphate-buffered saline (DPBS; 138 mM NaCl, 2.7 mM KCl, pH 7.4). Cell concentrations were determined using a Coulter Multisizer (Beckman Coulter).
6
Mycoplasma-free AML cell lines culture
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The mycoplasma-free AML U937 (ATCC CRL-1593.2; French-American-British/FAB phenotype M5, monoblast), NB4 (M3, promyelocyte) [65 (link)], HL-60 (ATCC 240-CCL; M2, myeloblast) and THP-1 (ATCC TIB-202; M5, monoblast) cells were cultured in complete RPMI 1640 medium supplemented with 5% FCS in a 5% CO2 humidified atmosphere at 37°C [66 (link)]. For every experiment, cells were harvested in log-phase proliferation at passage 16 or less. Cells (2 x 105/ml) were treated with IgG1 or anti-CD13 mAbs (10 μg/ml) for various periods of time. Cell proliferation was evaluated by counting the number of viable cells (with diameters ranging from 9 to 14 m) in a Coulter Multisizer (Beckman-Coulter, Villepinte, France).
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