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Home / Assay Kits

E. coli IspD Assay Kit Plus-100

SKU : BHE16300127

This product includes the reagents for 100 assays in a 384-well plate assay format including 0.400 ml of 10 x Buffer, 0.035 ml of 100 x MEP, 0.035 ml of 100 x CTP, 0.035 ml of 100 x E. coli IspD (200 nM), 0.035 ml of 100 x pyrophosphatase (PPase, 10 U/ml), and 5 ml of Dye for phosphate detection.

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Mfr.No.	ISPD100KE
Description	Methylerythritol phosphate cytidyltransferase (IspD) is one of the enzymes in the nonmevalonate pathway for isoprenoid biosynthesis that present in many pathogenic organisms and plants but absent in mammals. IspD is an attractive target for the development of novel antibiotics and herbicides. This enzyme catalyzes the formation of 4-diphosphocytidyl-2-C-methyl-D-erythritol from CTP and 2-C-methyl-D-erythritol 4-phosphate (MEP). The E. coli Methylerythritol Phosphate Cytidyltransferase (IspD) Assay is based on measurement of the pyrophosphate generated from the IspD reaction. The pyrophosphate is converted to phosphate by pyrophosphatase and the phosphate is detected by light absorbance at 650 nm. The assay is in a 384-well or 96-well plate format. It can be used for high throughput screening in drug discovery. The E. coli Methylerythritol Phosphate Cytidyltransferase (IspD) Assay Kit Plus-100 (Catalog # ISPD100KE) contains the reagents for 100 assays in a 384-well plate assay format including 400 µl of 10 x Buffer, 35 µl of 100 x MEP, 35 µl of 100 x CTP, 35 µl of 100 x E. coli IspD (200 nM), 35 µl of 100 x pyrophosphatase (PPase, 10 U/ml), and 5 ml of Dye MPA3000 for phosphate detection.
Drug target assays	Antibacterial drug targets

Product Manual	Datasheet
Protocols Description	ASSAY PROTOCOL The following assay protocol is based on the 384-well plate assay format. The reaction volume is 30 µl and the final assay volume is 75 µl. For 96-well plate assays, the reaction volume is 60 µl and the final assay volume is 150 µl. For detection using a cuvette, the reaction volume is 400 µl and the final assay volume is 1000 µl. 1. Reagent preparation: For each 10 assay reactions, (1) Prepare 297 µl of premix composed of 257.4 µl of H2O, 33 µl of 10 x Buffer, 3.3 µl of 100 x E. coli IspD and 3.3 µl of 100 x PPase . (2) Prepare 33 µl of 10 x Enzyme substrate by mixing 3.3µl of 100 x MEP, 3.3µl of 100 x CTP with 26.4µl of water. 2. Reaction: Mix 27 µl of the premix with 3 µl of the 10 x Enzyme substrate in each well. Incubate the reaction mixture at 37°C for 60 min. 3. Detection: Add 45 µl of the Dye MPA3000 into the 30 µl of the reaction mixture. Incubate for 5 min. Measure the light absorbance at 650 nm.

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Combination SKU	Supplier No.	Specifications	Size	Your price	Quantity
BHE16300127-1	ISPD100KE	enzyme included	100 assays	$613.14		Add to Cart
BHE16300127-2	ISPD500KE	enzyme included	500 assays	$1957.08		Add to Cart

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