phosphate assay

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Information about phosphate assay

Published on February 26, 2014

Author: teymourim901


PowerPoint Presentation: Phosphate assay remarks Presented by: Teymouri Manouchehr Theoretical mathematics v.s practical mathematics : Theoretical mathematics v.s practical mathematics We need to prepare a 2mg/ml Stock around 1 ml from 14.75 mg/ml Stock. How to conduct this? C1V1=C2V2 14.75*V1=2*1000µl V1=135.59 Error from rounding 14.75/2= 7.375 0.5932*7.375= 4.37 A practical solution 136*14.75=2* V2 V2 = 1003 It is just a simple math: It is just a simple math No change in the concentrations happens. No matter how many time you change your vessel Even throughout extruding procedure in extruding needles that you wash Phosphate assay is NOT a test to show you how much your phospholipid liquor is wasted N0. noooo teymouri you are absolutely wrong: N0. noooo teymouri you are absolutely wrong I have read in foreign article Nature says …… NO DISCUSSION PowerPoint Presentation: The concentration changes when the recruitment takes place. This includes proteins which tends to be denatured and attached to hydrophobic interfaces. even if this matters to so-called phospholipids waste, the phosphate assay is NOT conducted to calculate how much waste in phospholipids’ concentrations take place Then what phosphate assay is done for: Then what phosphate assay is done for It is conducted to show us what the total phospholipid concentration is after dialysis. Based on the test results, we can equal the phospholipid concentration of different formulations To show us how much Dxr is needed to fill the liposomes without any extra dxr left outside after active loading procedure PowerPoint Presentation: Formula 2 Formula 2 Formula 1 Formula 1 proceture: proceture Acid digestion Carbon deletion Start up kinetic reactio n Reading absorbance remarks: remarks Acid digestion: this causes any covalent bond in phospholipids breaks down H2O2: this is added to eliminate any carbon in the solution in a way that no color remains. Ascorbic acid and ammonium molibdate : both are added in excess to produce a blue color complex. Heat: to accelerate the endothermic reaction and in turn the color appearance. Molecula accessability: Molecula accessability PowerPoint Presentation: ABS= log [ABS%] 2=log[100] 47 47*2=94 47*3=141 47*4=188 47*5=235 46 46*2= 92 … 47 47*1.5=71 47*2=94 4.7*2.5=118 47*3=141 [P] stock kit=0.65 mM 0.65*47=30.55nM 0.65*94=61.1nM ABS ABS [P] nM [P] nM y= ax+c y= a’x+c Dialysis: Dialysis Reza and Ali are going to put their sample on dialysis with Sucrose 10%. Reza conduct the procedure three times, each time with 200 ml Sucrose 10%. Ali conduct the procedure 4 times. Each time with 100 ml Sucrose 10%. Who is getting the better dialysis efficiency? Dialysis: Dialysis Presume they have 1 ml formulation with 40000 mM of the component that they intended to get rid of. Dialysis efficiency = Dilution Reza procedure 40000/200=200 200/200=1 1/200= 0.005 Ali procedure 40000/100=400 400/100=4 4/100=0.04 0.04/100= 0.0004 0.0004 < 0.005 Dialysis: Dialysis I wana put my formulation on dialysis on the condition that my liposomal formulation at last surrounded in sucrose 10%, HEPES 10mM as well as the less HEPES consumed . How can I do that? first step= Sucrose 10% (100 ml) 40000/100= 400 Second step= Sucrose 10% (100 ml) 400/100= 4 Third Step= Sucrose 10% (100 ml) 4/100= 0.04 Forth step= Sucrose 10%, 10mM HEPES (100 ml) 0.04/100=0.0004 Then what about HEPES? Reverse dilution= 100 0.01 error then ultimate HEPES presence= 9.99 mM PowerPoint Presentation: Thank you

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