Primary HESCs cultured on chamber slides were transfected with either siRNA targeting HSPA8 or non-targeting siRNA and then either remained untreated or were decidualized with 8-Bromo-cAMP and MPA for 5 d. ER stress in control cultures was induced by treating cells with thapsigargin for 12 h (Sigma, UK). Cells were then fixed in 4% para-formaldehyde and permeabilized in 0.5% Triton. Primary antibodies, incubated overnight at 4°C in a humidified chamber (anti-LC3B 1:400). The secondary antibody used was labeled with Alexa Fluor 488 (1:200; Invitrogen). Slides were mounted with proGOLD (Invitrogen) and stained with 4′,6-diamidino-2-phenylindole (DAPI) to visualize nuclei. Images were captured using a Leica SP5 II confocal microscope.
Progold
Manufactured by Thermo Fisher Scientific
Sourced in Germany
ProGOLD is a laboratory equipment product designed for centrifugation. It provides high-speed separation of samples in a laboratory setting.
Automatically generated - may contain errors
Lab products found in correlation
Alexa fluor 488, by Thermo Fisher Scientific (2 mentions)
Sp5 2 confocal microscope, by Leica (1 mentions)
Thapsigargin, by Merck Group (1 mentions)
Lsm 710 laser scanning microscope, by Zeiss (1 mentions)
Alexa fluor 546, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
2 protocols using progold
1
Visualizing LC3B in Decidualized HESCs
2
Localization of NS1 and NUMA1 in Influenza-Infected A549 Cells
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A549 cells were grown overnight in Nunc 8-well Lab-Tek chamber slides. The next day, cells were treated with individual siRNA. After 48 h knockdown, cells were infected at MOI 3, incubated 20 h, and fixed with 3.7% formaldehyde for 20 min. Fixed cells were washed 4× with PBS, permeabilized with 0.2% Triton-X 100 for 10 min, washed again 4× with PBS, blocked with 3% BSA/PBS for 1 h and probed with monoclonal mouse α-NS1 [42 (link)] and with polyclonal rabbit α-NUMA1 (Bethyl Laboratory, A301-510A). Slides were washed 4 more times with PBS, then treated with Alexa Fluor® 546 (dilution of 1:250, Invitrogen)-conjugated goat anti-mouse secondary antibody, with Alexa Fluor® 488 (dilution of 1:250, Invitrogen)-conjugated goat anti-rabbit secondary antibody in 1% BSA/PBS for 1 h at room temperature, and with 4’, 6-diamidino-2-phenylindole (DAPI) (Invitrogen, dilution of 1:10000). Slides were washed three times with PBS, a drop of mounting media was added (Pro Gold, Invitrogen) to each spot and images obtained with a Zeiss LSM710 laser-scanning microscope (Carl Zeiss MicroImaging GmbH, Oberkochen, Germany), using 20× and 40× objectives.
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