Choline/Acetylcholine Quantification Colorimetric/Fluorometric Kit

Highly Sensitive Assay
Catalog #: K615 | abID: ab65345

Product Details

abID ab65345
Cat # +Size K615-100
Size 100 assays
Detection Method Absorbance (570 nm) or Fluorescence (Ex/Em 535/587 nm)
Species Reactivity Mammalian
Applications The kit can detect 10 pmol to ~5 nmol of choline or acetylcholine.
Features & Benefits • Simple procedure; takes ~40 minutes
• Fast and convenient
• Kit provides all necessary buffers and reagents for a accurate assay of Choline/Acetylcholine in various biological samples
Kit Components • Choline Assay Buffer
• Choline Probe
• Choline Enzyme Mix
• Acetylcholinesterase
• Choline Standard (5 µmol)
Storage Conditions -20°C
Shipping Conditions Gel Pack
USAGE For Research Use Only! Not For Use in Humans.


Choline and acetylcholine play important roles in many biological processes. BioVision’s Choline/Acetylcholine Quantification Kit provides a simple and sensitive means for quantifying Choline and Acetylcholine by either a colorimetric or fluorometric method. In the assay free choline is oxidized to betaine, via the intermediate betaine aldehyde. The reaction generates products which react with the Choline Probe to generate color (λ= 570 nm), and fluorescence (Ex/Em 535/587 nm). Acetylcholine can be converted to choline by adding acetylcholinesterase to the reaction. The kit can detect choline and acetylcholine (total choline – free choline) in various biological samples such as in blood, cells, culture media, fermentation media, etc. There is no need for pretreatment or purification of samples. The kit can detect 10 pmol~5 nmol of choline or acetylcholine.

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When using cell culture supernatant with K615, should we use choline-free media? Have you tested cell culture media?
We have not tested cell culture media with K615. However, studies show that acetylcholine is high in FBS. Therefore, it may be wise to use media without FBS for the assay. Reference: Lau et al (2012)and Lau et al (2013).
Do you recommend using hemolyzed serum with this kit? Is there any way to clean up these samples for the assay as we already bought 3 kits?
Hemolyzed samples are generally not recommended for any of BioVision’s enzymatic assays as it may interfere with the results. Therefore, we do not recommend using hemolyzed samples. However below are some recommendations for hemolyzed samples.

  a. Since Hemolyzed samples contain high amounts of iron, which may interfere with the assay and give a higher background, deproteinize the samples using 10 kDa filter.
   b. The end user can also add 1 mM of EDTA to their hemolyzed serum to chelate the iron.
How much cell lysate proteins should be used for this assay?
Typically we suggest using 100-200 ug for each assay. However, amount may vary among different tissues and cells depending on the Choline/Acetyl choline content.
Can milk powder be used as a sample for this kit?
Milk powder would have to dissolved homogeneously in the choline assay buffer (it is critical to ensure there are no particles or clumps) and then different dilutions can be prepared in the assay buffer and used for the assay.
Why is there under-estimation of choline content from standards spiked with serum samples?
There are serum proteins/enzymes that bind to choline/acetyl choline and hence it is possible that the recovery based on the calculations from this kit is lower than the expected value. Deproteinizing the serum sample could help.
The total choline ODs are the same as the free choline ODs, suggesting the ACh esterase is inactive. Why?
Possibilities for this outcome: 1. Exposure to many drugs can lead to serum inhibition of the acetylcholine esterase enzyme, especially in human samples. 2. There is extremely low levels of free acetyl choline in the samples (this is not unusual) and hence after addition of the esterase the total choline level is equal to the free choline.3. It would help to measure the background for the free and total choline so that after background subtraction the values can be compared.
Can protein content be used as an internal control for this assay?
Yes, a detergent-compatible BCA assay can be used for protein quantitation to normalize sample amount.
Are separate std curves recommended for choline and acetyl choline?
No, since during the detection reaction acetylcholine is converted to choline anyway.
Are trial sizes of this kit available?
Unfortunately, we do not have trial sizes of this kit available. However,for customers based in the US or Canada, a 10% off list price introductory discount can be offered on the list price. For all other customers based out of this area our regional BioVision distributor should be contacted.
What is the shelf life of this kit?
This kit is good for 12 months from the date of shipment in the unopened form when stored at the appropriate temperature and appropriate conditions. After opening and reconstitution, some of the components in this kit are good for 2 months at -20°C. Please refer to the datasheet for storage information and shelf life of each of the components.
Can frozen samples be used with this assay?
Fresh samples are always preferred over frozen samples. However, frozen samples can also be used, provided, they were frozen right after isolation, were not freeze thawed multiple time (for which we recommend aliquoting the samples before freezing) and have been frozen for relatively short periods.
Can individual components of this kit be purchased separately?
Yes, any of the kit's components can be purchased separately without having to buy the whole kit. Please refer to the component Cat #s mentioned on the datasheet for ordering.
Is it essential to make a standard curve for every expt, or is one curve/kit enough?
Yes, it is strongly recommended to do the standards every time you do the expt. There is always a chance that something was done differently that day and we do not want any conditions to differ between standards and samples.
How should I prepare my tissue sample for the assay ?
Choline Assay Buffer has detergent added to it and will help to lyse the tissue. Take 10 mg tissue and add 100 µl of Choline Assay Buffer. Homogenize the tissue using a dounce homogenizer or sonicator on ice (keep the tissue cold) and lyse the tissue. Once lysis is completed, centrifuge at 10,000g for 10 min. Take the supernatant and use it for the assay. You can use more than 10 mg tissue, but please increase the amount of Choline Assay Buffer proportionally to homogenize the tissue.
Contoli M et al., (2017) Up-Regulation of the Cholinergic System in a Model of Ex-Vivo Virus-Induced COPD Exacerbation - Role of Tiotropium Bromide, American Journal of Respiratory and Critical Care Medicine, 2017, 195:A6288
Ohshima et al., New Members of the Mammalian Glycerophosphodiester Phosphodiesterase Family: GDE4 AND GDE7 PRODUCE LYSOPHOSPHATIDIC ACID BY LYSOPHOSPHOLIPASE D ACTIVITY. J. Biol. Chem., Feb 2015; 290: 4260 - 4271.
Jamie K. Lau et al., Inhibition of Cholinergic Signaling Causes Apoptosis in Human Bronchioalveolar Carcinoma. Cancer Res., Feb 2013; 73: 1328 - 1339.
Su, X. et al. Requisite Role of the Cholinergic 7 Nicotinic Acetylcholine Receptor Pathway in Suppressing Gram-Negative Sepsis-Induced Acute Lung Inflammatory Injury. J. Immunol. 2010; 184: 401-410.
Liu et al., Protective Effects of Rosuvastatin in Experimental Renal Failure Rats via Improved Endothelial Function. Biol Res Nurs, Jul 2013; 15: 356 - 364.
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