A Homogenous, Universal Enzymatic Methyltransferase Assay for HMTs

AptaFluor – Leveraging the Power of Aptamers for Drug Discovery

Epigenetic regulation has been shown to be a contributing factor in a variety of diseases. The discovery of methyltransferase inhibitors is currently an area of intense activity. Methyltransferase enzymes methylate a variety of substrates using S-adenosylmethionine (SAM) as the methyl donor leaving the common product S-adenosylhomocysteine (SAH).

The AptaFluor SAH Methyltransferase Assay uses a natural occurring aptamer, or riboswitch, that selectively binds with SAH, the invariant product of methyltransferase reactions. The exquisite affinity and selectivity of the riboswitch combined with a positive TR-FRET signal enable screening and profiling of methyltransferases with unparalleled sensitivity.

By directly detecting SAH in a homogenous format, it is possible to perform screening and enzymatic studies with virtually any methyltransferase. The assay can be used as a tool to aid researchers in hit identification and characterization in a quest to develop new methyltransferase inhibitors as treatments for disease.

Direct Detection of SAH with a TR-FRET Readout

Ultra-Sensitive

The most sensitive HTS methyltransferase assay available with an LLD of 0.6 nM SAH. This dramatically reduces enzyme usage and allows the assay to be run at or below Km for SAM.

Compare Assay Sensitivity to Different Methods in this Poster

Use with Physiological SAM Concentrations Lower than 100nM

Stable

Outstanding reagent and signal stability reduces assay variability and allows for automated workflows.

Robust

Excellent Z’ under a variety of assay conditions.

PRMT4 enzyme with 0.2 uM or 2 uM SAM and histone substrate as positive controls. PRMT4 and histone substrate as negative control.

Simple Mix-and-Read

AptaFluor is in a simple, mix & read format. Run your enzyme reaction, add quenching reagent, add AptaFluor reagents, and read your plates.

Compatible with a Variety of MTase Acceptor Substrates

Toleration of MT acceptor substrates. 100 nM SAH-SAM standard curves were set up in the presence of nucleosomes (10 ng/μL), histone H3-3 (3 ng/μL), Histone H3 (1-21) peptide (10 μM) and Poly d (I-C) (2.5 mU/μL); control wells lacked a MT substrate.

Universal Assay – Use with Virtually Any Methyltransferase

Far Red TR-FRET Readouts Validated on Major Multimode Readers

Supplier Instrument TR-FRET Assays
bioteklogo TriStar²S LB 942 validated
Mithras² LB 943 validated
bioteklogo Cytation™ 5 validated
Cytation™ 3 validated
Cytation™ 1 validated
Synergy™ H1 validated
Synergy™ 2/H4/4 validated
Synergy™ HTX not capable
Synergy™ Neo 2 validated
BMGLABTECH Logo POLARstar® Omega validated
FLUOstar® Omega validated
PHERAstar® FSX validated
PHERAstar® Plus/FS validated
CLARIOstar® /Plus validated
MDS AT logo Analyst® GT/HT validated
Gemini® XPS/EM not capable
SpectraMax® M2/M2e not capable
SpectraMax® M5/M5e/FlexStation® 3 validated
SpectraMax® Paradigm contact us
perkinElmerLogo EnVision®/EnVision® Xcite validated
tecanLogo Infinite® M1000/M1000Pro/Safire2™ validated
Infinite® M200 not capable
Infinite® F500 validated
Infinite® F200/Ultraevolution contact us
Spark™ 10M validated

AptaFluor SAH Methyltransferase Assay Kits

* For custom or bulk orders (over 10,000 wells) please contact us (info@bellbrooklabs.com) for a quote.

A Guide to Navigating Hit Prioritization After Screening Using Biochemical Assays

So you have performed your screen. What’s next? This guide is focused on how biochemical assays are used for characterizing and prioritizing compounds following a primary screen with an enzyme target, whether using high throughput screening (HTS) or virtual screening (VS). A typical screening funnel is shown below, with the many applications of the biochemical activity assay highlighted.

Here we discuss strategies for hit-to-lead selection including:

  • Assay Considerations
  • Hit Confirmation
  • Running a Dose-Response
  • Compound Triaging
  • Hit Expansion
  • Mechanism of Action Studies
  • Residence Time Measurements
A Guide to Navigating Hit Prioritization After Screening Using Biochemical Assays

Enter your contact info and get some tips on hit triaging with this 9-page guide!

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