MCF-10A
Origin:
Schulze Lab
Cell Medium:
E4/F12+5%horse serum+20ng/mlEGF+100ng/mlCholera toxin+10ug/ml Insulin+0.5ug/mlHydrocortisone+PenStrep
Cell Seeding Density Per Well:
4000
Cell Culture Conditions:
10%CO2
Protocol:
Mix siRNA in 10uL HBSS to 37.5nM final concentration. Mix 0.1uL Lullaby in 10uL OPTIMEM. Mix together on plate and leave for 20 minutes. Add cells in 80ul/well. Change the media with 100ul fresh media after 24hours, then culture the cells for further 72hours
Delivery Reagent:
Lullaby
Delivery Reagent Cat No:
LL73000
Delivery Reagent Supplier:
Oz Bioscience
Delivery Reagent Concentration:
0.1ul/well
Silencing Reagent Concentration:
37.5nM
Silencing Reagent Supplier:
Dharmacon
Silencing Reagent Origin:
Schulze Lab
Description: Fixed in 80% Ethanol overnightFixation Compounds: EthanolCompound Concentration: 80%Treatment Time: long-term at -20C
Description: Cleaved caspase-3 staining
Blocking: 3%Donkey serum/PBS
Permeabilisation: 1hour
Primary Antibody - Cleaved caspase-3 (Asp175) antibody, Rabbit polyclonalSupplier: Cell Signalling (NEB)Cat No: 9661Batch No: Lot18Dilution: 1/750Incubation Time: 2hour
Secondary Antibody - Alexa Flour 488 donkey anti-rabbit IgG secondary antibodySupplier: Molecular ProbesCat No: A21206Batch No: 913921Dilution: 1/1500Incubation Time: 1 hour Nuclear Staining: DAPI
Nuclear Staining Concentration: 1ug/ml
Nuclear Staining Incubation Time: 1 hour
TTP Acumen Explorer Ch1 Localisation: nucleusStain: DAPITTP Acumen Explorer Ch2 Localisation: nucleusStain: Caspase-3