L-Lactate Assay Kit I

Sample Type:

Plasma, Serum, Cell culture supernatant, other body fluid

Method: Colorimetric method at 490nm

Sensitivity: 60µM-3000µM

Reaction Time: 30 minutes

Applications:

For biological research:

L-Lactate measurement in biological samples

For drug/pharm research:

Drug influence on L-Lactate metabolism

Storage: -80°C

Notes:

Materials needed but not supplied

A plate reader capable of measuring absorbance between
470-490nm

Adjustable pipettes and a repeat pipettor

A source of pure water; glass distilled water of HPLC-grade
water is sufficient

0.5M Acetic Acid

Clear flat bottom 96-well plates if not included in the kit
purchased

Shipping: Icepacks and/or Dry Ice

Reagent Preparation

1. L-Lactate
   Standards
   
   The vial contains 1000µl of 3mM L-Lactate Standard. The standard must be
   equilibrated to room temperature before use. 1ml of the standard is enough
   for making 3 standard curves if assayed in duplicate. Store at -80°C.
2. L-Lactate
   Assay Solution
   
   The solution contains enzymes that are light sensitive. The solution must
   be thawed on ice before use. Best to aliquot the amount needed and use it
   all to prevent thawing/freezing cycles. Freeze and store aliquots at -80°C.

Protocol

1. Sample Preparation

Serum/Plasma/other body fluid/cell culture supernatant

Serum, Plasma, other body fluid, or cell culture supernatant
can be measured directly by a series of dilutions of the sample (1/2; 1/4; 1/8;
.…..) to ensure the readings are within the standard curve. Your samples can be
diluted with dH2O.

Note: if samples (like hemolyzed serum/plasma or cell culture medium which
contain FBS) contain high level of lactate dehydrogenase capable of converting
lactate to pyruvate, it is important for the samples to be deproteinated.

Solid Samples

Solid samples, such as tissues, can be first homogenized and
extracted with ethanol (80%) with a tissues/Ethanol ratio of 1:8 (1 hr at 4°C)
followed by centrifugation at 10,000xg. The clear supernatants then can be
measured directly by a series of dilutions of the sample (1/2; 1/4; 1/8; .…..)
to ensure the readings are within the standard curve. Your samples can be
diluted with dH2O.

Adding Samples

Add 50μl of samples to each well. We recommend that samples
be assayed in duplicate.

Note: If prepared samples are not assayed the same day,
store the samples at -80°C. If samples need to be deproteinated, make sure to
deproteinize the samples prior to storing them in the freezer. The
deproteinated samples will be stable for one month while stored at -80°C. For
frozen samples, dilutions of samples must be done right before assaying.

2. Standard Curve Preparation

We recommend that L-Lactate Standards be assayed in
duplicate. A standard curve has to be run in each assay. Add 50µl, 40µl, 30µl,
20µl, 10µl, 5µl, 1 µl, and 0µl of L-lactate Standard to each well. Then adjust
volume to 50?l/well with dH2O.

3. Perform the assay

1. Add
   50ul of L-Lactate assay solution to each well containing L-Lactate
   standards and test samples.
2. Incubate
   for 30mins at 37°C incubator. Note: Please do not use CO2 incubator.
3. Stop
   the reaction by adding 50 ?l of 0.5 M acetic acid per well followed by
   brief gentle agitation. Note: Eliminate any air bubbles present in the
   wells using a needle prior to measurement.
4. Measure
   the absorbance at 490nM using a microplate reader.

4. Calculations

1. Average
   the OD490 nm values of replicate wells of each L-Lactate standard, test
   samples, and blank. In order to get the corrected absorbance, subtract the
   average OD490 nm value of the blank (L-Lactate Standard #8) from the
   average OD490 nm values from all standards and samples.
2. Make a
   standard curve by plotting OD490 nm values from each L-Lactate standards
   as a function of L-Lactate concentration. This can be done with excel
   spreadsheet. Calculate the value of L-Lactate in samples using the
   equation obtained from the linear regression of the standard curve.

L-Lactate(µM)= [ (Corrected absorbance)-(y-intercept)]/slope

Material Safety Data Sheet

Date: 09/30/25

1.       PRODUCT AND
COMPANY IDENTIFICATION

1.1    Product identifier

Product name: L-Lactate Assay Kit I

Catalog No: 1200011002, 120001100A, 120001100P,
1200012002, 120001200A, 12001200P, 1200014002, 120001400A, 120001400P

1.2    Relevant identified uses
of the substance or mixture and uses advised against

For research use only.

1.3    Details of the supplier of the
safety data sheet

Company: Abisara Bioscience, LLC.

Phone: 619-343-2616

1.4    Emergency telephone number

1-800-758-1630

2.       HAZARDS
IDENTIFICATION

2.1    Classification of the
substance or mixture

Not a hazardous substance or mixture

2.2    GHS Label elements, including
precautionary statements

Not a hazardous substance or mixture

2.3    Other hazards 

None

3.       COMPOSITION/INFORMATION
ON INGREDIENTS

3.1    Substances

L-lactate
Assay Solution

Components

CAS#

DMSO

67-68-5

L-LDH

9001-60-9

Tris-HCl,pH7.5

N/A

Water

7732-18-5

L-Lactate
Assay Standard

Components

CAS#

Sodium
L-Lactate

867-56-1

Tris-HCl,pH7.5

N/A

Water

7732-18-5

4.       FIRST AID
MEASURES

4.1    Description of first aid
measures

Eye contact

Check for and remove contact lenses and flush with copious
amounts of water; assure adequate flushing by separating the eyelids with
fingers; call a physician

Skin Contact

Flush with copious amounts of water; remove contaminated
clothing and shoes; call a physician

Inhalation

Remove to fresh air. If not breathing give
artificial respiration ;call a physician

Ingestion

If swallowed, never give anything by mouth to an unconscious
person. Rinse mouth with water ;call a physician

4.2    Most important symptoms and
effects, both acute and delayed

No information available

4.3    Indication of any immediate
medical attention and special treatment needed.

No information available

5.       FIRE FIGHTING
MEASURES

5.1    Extinguishing media

Suitable extinguishing media

Water spray, alcohol-resistant foam, dry chemical or carbon
dioxide

5.2    Special hazards arising from
the substance or mixture

No information available

5.3    Advice for firefighters

Wear self-contained breathing apparatus and protective
clothing to prevent contact with skin and eyes.

6.       ACCIDENTAL
RELEASE MEASURES

6.1    Personal precautions, protective
equipment and emergency procedures

Use full personal protective equipment. Evacuate personnel
to safe areas. Avoid breathing vapors, mist or gas.

6.2    Environmental precautions

Prevent further leakage or spillage. Prevent product from
entering drain.

6.3    Methods and material for
containment and cleaning up

Keep in suitable, closed containers for disposal

7.       HANDLING AND
STORAGE

7.1    Precautions for safe handling

Avoid skin/eye contact. Use protective equipment as needed.
Wash contaminated clothing before reuse.

7.2    Conditions for safe storage, including
any incompatibilities

Keep container tightly closed.  Store at -80°C.

7.3    Specific end use(s)

No data available.

8.       EXPOSURE
CONTROLS/PERSONAL INFORMATION

8.1    Control Parameters

This product contains no hazardous materials with
occupational exposure limits.

8.2    Exposure controls

Engineering controls

Provide shower and eye wash station

Personal protective equipment

Eye
Protection

Wear
safety goggles

Skin
Protection

Wear
protective clothing

Hand
Protection

Wear
protective gloves

Respiratory
protection

Wear
NIOSH/MSHA approved respirators

9.       PHYSICAL AND
CHEMICAL PROPERTIES

9.1    Information on basic physical
and chemical properties

L-Lactate Assay Solution

Appearance

Liquid,
yellowish

pH

7.5

Melting
point

No data
available

Freezing
point

No data
available

Boiling
point

No data
available

Density

No data
available

Water
solubility

Soluble

L-Lactate Standard

Appearance

Liquid,
colorless

pH

7.5

Melting
point

No data
available

Freezing
point

No data
available

Boiling
point

No data
available

Density

No data
available

Water
solubility

Soluble

9.2    Other safety information

No data available.

10.    STABILITY AND REACTIVITY

10.1Reactivity

No data available

10.2Chemical stability

Stable under recommended storage conditions.

10.3Possibility of hazardous reactions

No data available.

10.4Conditions to avoid

No data available

10.5Incompatible materials

Strong oxidizing agents

10.6Hazardous decomposition products

Hazardous decomposition products formed under fire
conditions: carbon oxides

11.    TOXICOLOGICAL INFORMATION

11.1Information on toxicological effects

Toxicological effects on this product have not been
thoroughly studied.

12.    ECOLOGICAL INFORMATION

12.1 Toxicity

Avoid release into environment

12.2 Persistence and degradability

No data available

12.3 Bioaccumulative potential

No data available

12.4 Mobility in soil

No data available

12.5 Results of PBT and vPvB assessment

No data available

12.6 Other adverse effects

No data available 

13. DISPOSAL INFORMATION

13.1 Waste treatment methods

Dispose in accordance with local, state, and federal
regulations.

14. TRANSPORT INFORMATION

DOT:

Proper shipping name: none

Non-Hazardous for transport: this substance is considered to be non-hazardous
for transport

IATA:

Proper shipping name: none

Non-Hazardous for transport: this substance is considered to be non-hazardous
for transport

ADR/RID

Proper shipping name: none

Non-Hazardous for transport: this substance is considered to be non-hazardous
for transport

 15.    REGULATORY INFORMATION

EU Risk and Safety phrases:

R36/37/38: irritating to eyes/respiratory system/skin

 16.    OTHER INFORMATION

The information above is believed to be accurate and
represents the best information currently available to us. However, we make no
warranty of merchantability or any other warranty, express or implied, with
respect to such information. Abisara Bioscience, LLC. shall not be held liable
for any damages or other consequences resulting from handling or from contact
with the above product.

Q. What type of medium should I use for making cultured
cells for Lactate assay?

A. Please use phenol red free medium. Please do not use
phenol red medium since phenol red would affect absorbance readings. 

Q. What enzymes is used in Lactate Assay?

A. It is LDH.

Q. Can you tell me the compositions of your Lactate
Assay?

A. Unfortunately it is proprietary.

Q. How do I know whether I need to use L-Lactate or
D-Lactate assay kit?

A. L-Lactate assay kits are commonly used for measuring
lactate level in animal cells.

D-lactate assay kits are commonly used for measuring lactate
level in bacteria cells. 

Q. Do I need to make a standard curve everytime?

A. Yes, it is necessary since you calculate your sample
concentration based on the standard curve. Please do not use the
representative Lactate standard curve in the protocol to calculate
concentrations of your samples.

Q. Can I use fluorescence spectroscopy to measure
readings for Lactate assay kits?

A. No, the kit does not work with a fluorescence reader
since the kit employs colorimetric assays.

Q. How do I measure Lactate from cultured cells?

A. You can grow cells in phenol red-free medium, then you
can extract cells with 80% ethanol.

Q. Can I use frozen samples?

A. Although it is better to use fresh samples for assays,
you can use frozen samples. If you are not using frozen samples in once,
please make aliquots of your samples before you put them in a freezer to
prevent from degrading samples from repeated freeze-thaw cycles.

Q. My sample formed preciptation after adding acetic
acid. How can I prevent this?

A. You can still measure readings without adding acetic
acid.

 Q. The protocol recommends measuring the absorbance
at 490nm. Is 485nm close enough?

A. It is ok as long as you set up the absorbance between 470
and 490nm.

Q. Will L-Lactate assay measure D-Lactate?

A. No, it wont.

Q. Can I use a transparent wall 96-well plate with
Lactate Kit?

A. Yes, you can.

Q. Can Ethanol interfere with the enzyme?

A. The enzyme can tolerate up to 20% of ethanol without
affecting it’s enzymatic activity with L-Lactate.

Sam Van de Velde, Meghan F. Hogan, and Marc Montminy.:mTOR
links incretin signaling to HIF induction in pancreatic beta cells.PNAS, Oct
2011; 108: 16876 - 16882.

Jun Mifune, Katrin Grage, and Bernd H. A. Rehm.:Production of Functionalized
Biopolyester Granules by Recombinant Lactococcus lactis.Appl. Envir.
Microbiol., Jul 2009; 75: 4668 - 4675.

Dragutin J. Savic, William M. McShan, and William M. McShan.:Long-term survival
of Streptococcus pyogenes in rich media is pH-dependent.Microbiology, Jun 2012;
158: 1428 - 1436

Kristin A. Anderson, Fumin Lin, Thomas J. Ribar, Robert D. Stevens, Michael J.
Muehlbauer, Christopher B. Newgard, and Anthony R. Means.:Deletion of CaMKK2
from the Liver Lowers Blood Glucose and Improves Whole-Body Glucose Tolerance
in the Mouse.Mol. Endocrinol., Feb 2012; 26: 281 - 291.

Janina P. Lewis, Divya Iyer, and Cecilia Anaya-Bergman.:Adaptation of
Porphyromonas gingivalis to microaerophilic conditions involves increased
consumption of formate and reduced utilization of lactate.Microbiology, Nov
2009; 155: 3758 - 3774.

Amparo Wolf, Sameer Agnihotri, Johann Micallef, Joydeep Mukherjee, Nesrin
Sabha, Rob Cairns, Cynthia Hawkins, and Abhijit Guha.:Hexokinase 2 is a key
mediator of aerobic glycolysis and promotes tumor growth in human glioblastoma
multiforme.J. Exp. Med., Feb 2011; 208: 313 - 326.

Inna Serganova, Asif Rizwan, Xiaohui Ni, Sunitha B. Thakur, Jelena Vider, James
Russell, Ronald Blasberg, and Jason A. Koutcher.:Metabolic Imaging: A Link
between Lactate Dehydrogenase A, Lactate, and Tumor Phenotype.Clin. Cancer
Res., Oct 2011; 17: 6250 - 6261.

Changlu Liu, Jiejun Wu, Jessica Zhu, Chester Kuei, Jingxue Yu, Jonathan
Shelton, Steven W. Sutton, Xiaorong Li, Su Jin Yun, Taraneh Mirzadegan, Curt
Mazur, Fredrik Kamme, and Timothy W. Lovenberg.:METABOLISM, REGULATION, AND
SIGNALING: Lactate Inhibits Lipolysis in Fat Cells through Activation of an
Orphan G-protein-coupled Receptor, GPR81.J. Biol. Chem., Jan 2009; 284: 2811 -
2822.

Marina Ostroukhova, Nicholas Goplen, Md Zunayet Karim, Lidia Michalec, Lei Guo,
Qiaoling Liang, and Rafeul Alam.:The role of low-level lactate production in
airway inflammation in asthma.Am J Physiol Lung Cell Mol Physiol, Feb 2012;
302: L300 - L307.

Zhuang, Yongxian, Daniel K Chan, and W Keith Miskimins.
“Preventing Feedback Activation of Glycolytic ATP Production Enhances Metformin
Cytotoxicity in Breast Cancer Cells When Oxidative Phosphorylation Is
Inhibited.” Cancer & Metabolism 2.Suppl 1 (2014): P89. PMC. Web. 6 Oct.
2015.

Lezi, E. et al. “Effect of Exercise on Mouse Liver and Brain
Bioenergetic Infrastructures.” Experimental physiology 98.1 (2013): 207–219.
PMC. Web. 6 Oct. 2015.

Yongxian Zhuang,Daniel K.Chan,Allison B.Haugrud,W.Keith
Miskimins. Mechanisms by Which Low Glucose Enhances the Cytotoxicity of
Metformin to Cancer Cells Both In Vitro and In Vivo. September 25, 2014 DOI:
 10.1371/journal.pone.0108444

Borges FT1, Melo SA, Özdemir BC, Kato N, Revuelta I, Miller
CA, Gattone VH 2nd, LeBleu VS, Kalluri R. TGF-?1-containing exosomes from
injured epithelial cells activate fibroblasts to initiate tissue regenerative
responses and fibrosis. J Am Soc Nephrol. 2013 Feb;24(3):385-92. doi:
10.1681/ASN.2012101031. Epub 2012 Dec 28.