50 μl of apical and 50 μl basolateral cell culture supernatants were analyzed for cytokines using the Meso Scale Discovery (MSD) V-plex Viral Panel 1 Human Kit (Meso Scale Diagnostics, Rockville, MA) as per the manufacturer’s instructions. Briefly, 1:5 dilutions of cell supernatant samples were diluted in PBS containing 1% Triton-X. Samples were added to the MSD plate along with a 7-point 4-fold serial dilution (concentrations related to certificate of analysis for each individual standard) of protein standards diluted in PBS with 1% Triton-X. The MSD plate was sealed, and samples incubated at room temperature for 2 hours on a plate shaker (ThermoFisher Scientific, Waltham, MA) at 700RPM. The plate was washed 3x in wash buffer and 25 μl of secondary antibody was added to each well. Plates were sealed and incubated at room temperature on a plate shaker at 700RPM for a further 2 hours in the dark. Plates were washed 3x with wash buffer and 50 μl of 2x read buffer (MSD R92TC) was added to each well. The plates were read on the MESO Sector S 600 (Meso Scale Diagnostics) and concentrations determined against the standard curves.
Plate shaker
Manufactured by Thermo Fisher Scientific
The Plate Shaker is a laboratory instrument designed to agitate and mix samples in microplate or tube formats. It provides consistent and controlled orbital motion to ensure thorough mixing of the contents. The core function of the Plate Shaker is to facilitate efficient sample preparation and homogenization in a variety of laboratory applications.
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Lab products found in correlation
2 protocols using plate shaker
1
Cytokine Quantification from Cell Supernatants
2
MSD Multiplex Cytokine Quantification
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50 μl of cell culture supernatants were analyzed for cytokines using the Meso Scale Discovery (MSD) Proinflammatory panel 1, Human kit, Lot:K0081459 & K0081080 (Meso Scale Diagnostics, Rockville, MD) as per the manufacturer’s instructions. Briefly, 1:5 dilutions of cell supernatant samples were diluted in PBS containing 1% Triton-X. Samples were added to the MSD plate along with a 7-point 4-fold serial dilution (concentrations related to certificate of analysis for each individual standard) of protein standards diluted in PBS with 1% Triton-X. The MSD plate was sealed, and samples incubated at room temperature for 2 hours on a plate shaker (ThermoFisher Scientific) at 700RPM. The plate was washed 3x in wash buffer and 25 μl of secondary antibody was added to each well. Plates were sealed and incubated at room temperature on a plate shaker at 700RPM for a further 2 hours in the dark. Plates were washed 3x with wash buffer and 50 μl of 2x read buffer (MSD R92TC) was added to each well. The plates were read on the MESO Sector S 600 (Meso Scale Diagnostics), and concentrations determined against the standard curves.
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