Cytomix Buffer
10 mmol/L K2HPO4/KH2PO4(pH 7.6)、120 mmol/L KCl、0.15 mmol/L CaCl2、5 mmol/L MgCl2、25 mmol/L Hepes(pH 7.6)、2 mmol/L EDTA，adjust with 10 mol/L KOH to pH 7.6，then filter with 0.22 μm filter and alquoted.
Dear Andreia Ribeiro Albuquerque,
The Giardia trophozoites (3 3 106 cells) were examined
microscopically, pelleted, and suspended in 50 ml of luciferase lysis buffer
(100 mM K2HPO4-KH2PO4 [pH 7.8], 1 mM dithiothreitol, 2 mM EDTA, 1%
Triton X-100) containing 75 mM trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane
(E-64; Sigma) and 0.75 mg of leupeptin (Sigma) per ml. Samples were
immediately frozen at 2708C for more than 1 h and thawed on ice for 10 min. After
warming to room temperature, 10 ml of the cell lysate was assayed in 100 ml of
luciferase assay reagent (30 mM Tris-HCl [pH 7.8], 3 mM ATP, 15 mM MgSO4, 10
mM dithiothreitol, 0.2% Triton X-100) by injecting 100 ml of 0.5 mM D-luciferin
(Sigma) to initiate the reaction according to the manufacturer’s instructions. The
data thus obtained were in relative light units as measured with a luminometer
(Monolight 2010; Analytical Luminescence Laboratory, San Diego, Calif.)
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC230732/pdf/154867.pdf
Hello to all and thank you for your answers. I believe the correct reagent for the cytomix, among various is the K2HPO4/KH2PO4 mix and not KHPO4, probably the latter being a typo from an old protocol.
Also strugling with finding the correct GSH (Glutathione), since there are several available (oxidised, reduced...) I wonder what would be the correct one for transfection and what would be its "function" in the cytomix...
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