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CS0340

Dihydrofolate Reductase Assay Kit

1 kit sufficient for 50-100 tests

Synonym(s):

DHFR Assay Kit

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About This Item

NACRES:
NA.84
UNSPSC Code:
12161503
Pricing and availability is not currently available.
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Quality Segment

200

usage

 kit sufficient for 50-100 tests

shipped in

dry ice

storage temp.

−20°C

Gene Information

human ... DHFR(1719)

Application

The kit is used for the detection of DHFR activity and screening of DHFR inhibitors. The assay is based on the ability of DHFR to catalyze the reversible NADPH-dependent reduction of dihydrofolic acid to tetrahydrofolic acid. The reaction progress is followed by monitoring the decrease in absorbance at 340 nm.
Dihydrofolic acid+NADPH+H+ ↔ Tetrahydrofolic acid+NADP+

Biochem/physiol Actions

DHFR (dihydrofolate reductase) is an enzyme that catalyzes a reaction essential for the biosynthesis of nucleotidic bases of DNA. Blocking the enzyme causes cell death as a result of DNA synthesis inhibition. DHFR is an excellent target for anti-tumor drugs.

Features and Benefits

  • Quick and simple method.
  • The kit contains all the reagents required for a colorimetric assay of DHFR activity in cell lysates, tissue homogenates, or column fractions of purified enzyme.
  • The kit includes a purified enzyme for use as a positive control and screening of DHFR inhibitors.
  • The kit includes methotrexate (MTX), a prokaryotic and eukaryotic DHFR specific inhibitor, which exhibits anti-tumor activities.
  • The kit was tested on A431, NIH-3T3, and CHO cell lines, rat liver, kidney, brain, and skeletal muscle tissue extracts, and recombinant DHFR.

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Dihydrofolate Reductase Assay Kit 1 kit sufficient for 50-100 tests

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Gene Information

human ... DHFR(1719)

Gene Information

human ... HDAC1(3065)

Gene Information

human ... HMGCR(3156)

Gene Information

human ... TGM1(7051), TGM2(7052), TGM3(7053), TGM4(7047), TGM5(9333), TGM6(343641), TGM7(116179)

Quality Level

200

Quality Level

200

Quality Level

200

Quality Level

200

usage

 kit sufficient for 50-100 tests

usage

sufficient for 100 assays (96 well plates)

usage

sufficient for 100 assays (200 μL), sufficient for 30 assays (1 mL)

usage

-

storage temp.

−20°C

storage temp.

−20°C

storage temp.

−70°C

storage temp.

−20°C

shipped in

dry ice

shipped in

dry ice

shipped in

dry ice

shipped in

dry ice

Kit Components Only

Product No.
Description
  • Assay Buffer 10x for DHFR 30 mL
  • Dihydrofolate Reductase (DHFR) human .1 U
  • Dihydrofolic acid (DHFR substrate) 3 x 10
  • Amethopterin (+)(methotrexate, MTX)
    (DHFR inhibitor) 2 x 10
  • NADPH (β-Nicotinamide adenine dinucleotide phosphate reduced tetrasodium salt) 25 mg

pictograms

Skull and crossbonesHealth hazard
GHS06,GHS08

signalword

Danger

Storage Class

6.1C - Combustible acute toxic Cat.3 / toxic compounds or compounds which causing chronic effects

flash_point_f

Not applicable

flash_point_c

Not applicable

wgk

WGK 3

Hazard Classifications

Acute Tox. 3 Oral - Eye Irrit. 2 - Repr. 1B - Skin Irrit. 2 - STOT SE 2 - STOT SE 3

target_organs

Eyes,Central nervous system, Respiratory system

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Certificates of Analysis (COA)

Lot/Batch Number
0000524266
0000524266
0000381587
0000306369
0000381587

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Global Trade Item Number

SKUGTIN
CS0340-1KT04061833553084

Questions

1–2 of 2 Questions  

  1. 1. what is the origin of this DHFR. Is it from Human, E. coli or any other. 2. I want to relate antibacterial activity of my compound with this enzyme assay kit. Is it obvious.

    1 answer

    1. The Dihydrofolate Reductase that is provided in the kit is product D6566, a recombinant human enzyme that is expressed in E. coli. Please see the link below to review the datasheet for this component:
      https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/213/424/d6566dat.pdf

      One intended use of this kit is for screening DHFR inhibitors, including inhibitors of bacterial DHFR. This kit is expected to be suitable for the measurement of antibacterial activity. This characteristic is expressed as DHFR inhibitory activity of a compound of interest. Please see the link below to review the datasheet for this kit:
      https://www.sigmaaldrich.com/deepweb/assets/sigmaaldrich/product/documents/427/408/cs0340bul.pdf

      Helpful?

  2. I want to measure activity in protein variants that have different cellular concentration (protein stability), will this affect my activity?

    1 answer

    1. It is unclear the relation between cellular concentration and protein stability. It seems the intention is to produce dihydrofolate reductase recombinantly and then measure the activity. Protein stability is dependent on several different factors such as pH, temperature, buffering conditions, storage, purification, etc. If a protein is not stable it can certainly affect the activity. Testing would be necessary to determine the overall impact on the activity as you optimize your process.

      To further discuss this application, we kindly ask you to navigate to the link https://www.sigmaaldrich.com/techservice, and click on "Product Technical Inquires" under the Products Section with all the required information so that a member of our team can reach out to you to assist further. Thank you.

      Helpful?

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