In the past, measurement of SIRT1 activity is primarily based on the work with radioactive materials (radioactive acetylated histone as a substrate). Recently, the most used method for measuring the activity of deacetylase without the use of radioactive substances is fluorimetric. The principle is based on changing a SIRT1 reaction into the activity of the protease.

The kits usually have different reactivity (it depends on what species are you working with), detection range and price. I think that SIRT1 Activity Assay Kit (Fluorometric) (ab156065) (Abcam) is a good choice. If you are working with human cells SIRT1 Assay Kit (Sigma) also would be suitable.

You can find here some of the available kits in the market:

SIRT1 Activity Assay Kit (Fluorometric) (ab156065) (Abcam)

http://www.abcam.com/sirt1-activity-assay-kit-fluorometric-ab156065.html

SIRT1 Assay Kit (Sigma)

https://www.sigmaaldrich.com/catalog/product/sigma/cs1040?lang=en®ion=BG

Sirtuin Activity Assay Kit (Fluorometric) (Biovision)

https://www.biovision.com/sirtuin-activity-assay-kit-fluorometric.html

SIRT1 Direct Fluorescent Screening Assay Kit (Cayman Chemical)

https://www.caymanchem.com/product/10010401

SIRT1 (Sirtuin1) Fluorogenic Assay Kit (BPS Biosciece)

http://bpsbioscience.com/fluorogenic-sirt1-sirtuin1-assay-kit-50081

In the past, measurement of SIRT1 activity is primarily based on the work with radioactive materials (radioactive acetylated histone as a substrate). Recently, the most used method for measuring the activity of deacetylase without the use of radioactive substances is fluorimetric. The principle is based on changing a SIRT1 reaction into the activity of the protease.

The kits usually have different reactivity (it depends on what species are you working with), detection range and price. I think that SIRT1 Activity Assay Kit (Fluorometric) (ab156065) (Abcam) is a good choice. If you are working with human cells SIRT1 Assay Kit (Sigma) also would be suitable.

You can find here some of the available kits in the market:

SIRT1 Activity Assay Kit (Fluorometric) (ab156065) (Abcam)

http://www.abcam.com/sirt1-activity-assay-kit-fluorometric-ab156065.html

SIRT1 Assay Kit (Sigma)

https://www.sigmaaldrich.com/catalog/product/sigma/cs1040?lang=en®ion=BG

Sirtuin Activity Assay Kit (Fluorometric) (Biovision)

https://www.biovision.com/sirtuin-activity-assay-kit-fluorometric.html

SIRT1 Direct Fluorescent Screening Assay Kit (Cayman Chemical)

https://www.caymanchem.com/product/10010401

SIRT1 (Sirtuin1) Fluorogenic Assay Kit (BPS Biosciece)

http://bpsbioscience.com/fluorogenic-sirt1-sirtuin1-assay-kit-50081

Dimitar Parvanov: Thank you for the suggestions. I will certainly look into them as options.

Hi everyone.

I am considering to buy SIRT1 Activity Assay Kit (Fluorometric) (Abcam cat. no. ab156065). However, I read very carefully the protocol booklet, and I have a strong doubt. I am looking for a method that detects ONLY the activity of SIRT1. Does the kit eliminate the contribution of other sirtuins by measuring sirtuin-dependent deacetylase activity on proteins derived from isolated nuclei? How can Abcam be sure that the kit does not detect the activity of other nuclear sirtuins (SIRT2/6/7)? Unfortunately, the datasheet of product no. ab156065 provides no information with regard to the substrate peptide used. Does the kit use a substrate peptide that is very specific for SIRT1? Does anybody know which one?

I've just sent a similar email to the Abcam's technical support.

Thanks in advance.

Stefano Falone Department of Life, Health and Environmental Sciences University of L'Aquila Email [email protected]

Stefano Falone, I have a similar concern about Sirt1 Activity Assay (Abcam cat. no. ab156065), could you be able to get an answer for your doubts as well? I am currently using Universal Sirt Activity Assay (Abcam, ab156915) which measures all deacetylated lysines by all isoforms of sirtuins colorimetrically. Since I am using nuclear and cytoplasmic fractions of the human cancer cells and since sirtuin isoforms distributed differently in these fractions, according to the kit's results, I may discuss the activity of specific isoform of sirtuins, however, it is not a reliable and clear cut way. That is why I should use isoform-specific activity assays as well but I have concerns about their specificity. If you could help, it would be very nice.

Büşra Binarci , I faced this problem in 2019. Then, we decided to immunoprecipitate SIRT1, first. In order to do this, we used protein A agarose beads and anti-SIRT1 antibody-conjugate (cat. ab193254 and ab7343, Abcam, respectively). Briefly, the agarose beads were mixed with primary antibody for 4h at 4 °C (15µg of anti-SIRT1 antibody/mL of beads). The mixture were then centrifuged and washed with the lysis buffer (20 mM Tris-HCl, pH 7.4, 135 mM NaCl, 1.5 mM MgCl2, 1 mM EGTA, 10% glycerol, 1% Nonidet P-40) several times. After the last wash, cell lysates (2 mg of total proteins) were incubated overnight with the antibody-beads mixture. The immunecomplexes were then centrifuged and washed, and the immunoprecipitates were directly used for the SIRT1 activity assay kit (cat. ab156065, Abcam), according to supplier’s instructions.

You can find this method described in our paper, which was published onto Antioxidants (DOI: 10.3390/antiox8090346).

I hope this helps you.

Best,

Stefano

Thank you so much Stefano Falone