Lactate Assay Kit

N° du produit ABIN1000300

Détail du produit

Antigène: Lactate

Application: Biochemical Assay (BCA)

Type d'échantillon: Serum, Plasma, Cell Culture Supernatant

Specificité: 0.05 mM

Attributs du produit: Sensitive and accurate. Detection limit of 0.05 mM and linearity up to 2 mM L-Lactate in 96-well plate assay. For cell culture samples containing phenol red: detection limit of 0.1 mM and linearity up to 1 mM L-Lactate in 96-well plate assay.
Convenient. The procedure involves adding a single working reagent, and reading the optical density at time zero and at 20 min. Room temperature assay. No 37°C heater is needed.
High-throughput. Can be readily automated as a high-throughput 96- well plate assay for thousands of samples per day.

Ingrédients: Assay Buffer: 10 mL. NAD Solution: 1 mL. Enzyme A: 120 µL. MTT Solution: 1.5 mL. Enzyme B: 120 µL. Standard: 1.0 mL 20 mM L-Lactate.

Matériel non inclus: Pipeting (multi-channel) devices. Clear-bottom 96-well plates (e.g. Corning Costar) and plate reader.

Information d'application

Indications d'application: Direct Assays: lactate in serum, plasma, and cell media samples.

Commentaires:

The following substances interfere and should be avoided in sample preparation: ascorbic acid, SDS (>0.2%), sodium azide, NP-40 (>1%) and Tween-20 (>1%). 0.0 0.5 1.0 1.5 2.0 0.0 0.2 0.4 0.6 L-Lactate [L-Lactate](mM) OD565 Standard Curve in 96-well plate assay in water.

Protocole: Important: this assay is based on an enzyme-catalyzed kinetic reaction. Addition of Working Reagent should be quick and mixing should be brief but thorough. Use of a multi-channel pipettor is recommended.
1. Standard Curve. Prepare 1000 µL 2.0 mM L-lactate Premix by mixing 100 µL 20 mM Standard and 900 µL distilled water. For cell culture samples containing phenol red, prepare 1000 µL >1.0 mM lactate Premix by mixing 50 µL 20 mM Standard and 950 µL culture medium without serum. Transfer 20 µL standards into wells of a clear bottom 96-well plate. Samples. Add 20 µL sample per well in separate wells. For samples with potential endogenous enzyme activity (i.e. serum, plasma, tissue extracts), two reactions should be run: one with added Enzyme A and a No Enzyme A control. Serum and Plasma should be diluted at least 2° with dH2O prior to the assay.
2. Reagent Preparation. Spin the Enzyme tubes briefly before pipetting. For each Sample and Standard well, prepare Working Reagent by mixing 60 µL Assay Buffer, 1 µL Enzyme A, 1 µL Enzyme B, 10 µL NAD and 14 µL MTT. Fresh reconstitution is recommended. For the No Enzyme A sample control, the Working Reagent includes 60 µL Assay Buffer, 1 µL Enzyme B, 10 µL NAD and 14 µL MTT.
3. Reaction. Add 80 µL Working Reagent per reaction well quickly. Tap plate to mix briefly and thoroughly.
4. Read optical density (OD0) for time zero at 565 nm (520-600nm) and OD20 after a 20-min incubation at room temperature.
5. Calculation. Subtract OD0 from OD20 for the standard and sample wells. Use the OD values to determine the sample L-lactate concentration from the standard curve. For samples requiring a No Enzyme A control, subtract the ODNoEnz value from the ODSample and use this OD value to determine the sample L-lactate concentration from the standard curve. Note: if the sample OD value is higher than OD for 2 mM L-lactate standard, dilute sample in water and repeat the assay. Multiply the results by the dilution factor.

Restrictions: For Research Use only

Stockage

Stock: -20 °C

Référence

Wang, Li, Mao, Li, Liu, Chen, Lv, Wang, Wu, Dai, Wang, Zhao, Tang, Sun: "Ras-induced epigenetic inactivation of the RRAD (Ras-related associated with diabetes) gene promotes glucose uptake in a human ovarian cancer model." dans: The Journal of biological chemistry, Vol. 289, Issue 20, pp. 14225-38, (2014) (PubMed).

Le Nihouannen, Barralet, Fong, Komarova: "Ascorbic acid accelerates osteoclast formation and death." dans: Bone, Vol. 46, Issue 5, pp. 1336-43, (2010) (PubMed).

Fan, Dickman, Zong: "Akt and c-Myc differentially activate cellular metabolic programs and prime cells to bioenergetic inhibition." dans: The Journal of biological chemistry, Vol. 285, Issue 10, pp. 7324-33, (2010) (PubMed).

Krastel, Senadheera, Mair, Downey, Goodman, Cvitkovitch: "Characterization of a glutamate transporter operon, glnQHMP, in Streptococcus mutans and its role in acid tolerance." dans: Journal of bacteriology, Vol. 192, Issue 4, pp. 984-93, (2010) (PubMed).

Toschi, Lee, Thompson, Gadir, Yellen, Drain, Ohh, Foster: "Phospholipase D-mTOR requirement for the Warburg effect in human cancer cells." dans: Cancer letters, Vol. 299, Issue 1, pp. 72-9, (2010) (PubMed).

Wei, Kulp, Chen: "Energy restriction as an antitumor target of thiazolidinediones." dans: The Journal of biological chemistry, Vol. 285, Issue 13, pp. 9780-91, (2010) (PubMed).

Milovanova, Bhopale, Sorokina, Moore, Hunt, Hauer-Jensen, Velazquez, Thom: "Hyperbaric oxygen stimulates vasculogenic stem cell growth and differentiation in vivo." dans: Journal of applied physiology (Bethesda, Md. : 1985), Vol. 106, Issue 2, pp. 711-28, (2009) (PubMed).

Devalaraja-Narashimha, Padanilam: "PARP-1 inhibits glycolysis in ischemic kidneys." dans: Journal of the American Society of Nephrology : JASN, Vol. 20, Issue 1, pp. 95-103, (2009) (PubMed).

Kashimshetty, Desai, Kale, Lee, Moland, Branham, New, Chan, Younis, Boelsterli: "Underlying mitochondrial dysfunction triggers flutamide-induced oxidative liver injury in a mouse model of idiosyncratic drug toxicity." dans: Toxicology and applied pharmacology, Vol. 238, Issue 2, pp. 150-9, (2009) (PubMed).

Weber, Mitchell, Veliotes, Mitchell, Kamerman: "Hyperalgesia induced by oral stavudine administration to rats does not depend on spinal neuronal cell death, or on spinal or systemic inflammatory cytokine secretion, or metabolic dysregulation." dans: Neurotoxicology, Vol. 30, Issue 3, pp. 423-9, (2009) (PubMed).

Roy, De, Mukherjee, Chander, Konar, Bandyopadhyay, Bandyopadhyay: "Excess of glucocorticoid induces cardiac dysfunction via activating angiotensin II pathway." dans: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology, Vol. 24, Issue 1-2, pp. 1-10, (2009) (PubMed).

Aronoff, Lewis, Serezani, Eaton, Goel, Phipps, Peters-Golden, Mancuso: "E-prostanoid 3 receptor deletion improves pulmonary host defense and protects mice from death in severe Streptococcus pneumoniae infection." dans: Journal of immunology (Baltimore, Md. : 1950), Vol. 183, Issue 4, pp. 2642-9, (2009) (PubMed).

Senadheera, Krastel, Mair, Persadmehr, Abranches, Burne, Cvitkovitch: "Inactivation of VicK affects acid production and acid survival of Streptococcus mutans." dans: Journal of bacteriology, Vol. 191, Issue 20, pp. 6415-24, (2009) (PubMed).

Zheng, Delgoffe, Meyer, Chan, Powell: "Anergic T cells are metabolically anergic." dans: Journal of immunology (Baltimore, Md. : 1950), Vol. 183, Issue 10, pp. 6095-101, (2009) (PubMed).

Détail du antigène

Antigène: Lactate

Abstract: Lactate Produits

Sujet: Quantitative determination of L-lactic acid by colorimetric (565nm) method.
Procedure: 20 min.

Lactate is generated by lactate dehydrogenase (LDH) under hypoxic or anaerobic conditions. Monitoring lactate levels is, therefore, a good indicator of the balance between tissue oxygen demand and utilization and is useful when studying cellular and animal physiology. Simple, direct and automation-ready procedures for measuring lactate concentration are very desirable. This lactate assay kit is based on lactate dehydrogenase catalyzed oxidation of lactate, in which the formed NADH reduces a formazan (MTT) reagent. The intensity of the product color, measured at 565 nm, is proportionate to the lactate concentration in the sample.