The best thing about Montana Molecular’s biosensors has been the ease of use. My lab has already been doing live cell imaging, mostly free Ca2+. All we had to do is to infect the cells and use the same microscope set-up to gather data on PIP2, DAG and cAMP.Vladlen Z Slepak, PhD
Robust Assays in Living Cells
» Easy assay steps – No cell lysis
» BacMam delivery to most cell types
» Fluorescence imaging or plate reader detection (Z’ > 0.8)
» Physiologically-relevant and cell friendly
» No forskolin or IBMX needed
» No enzymes or co-factors
» Multiplex GPCR Assays: Gs, Gq, Gi, Voltage
Simultaneous measurements of GPCR signaling in live HEK 293 cells. Green sensors in the top panels, red sensors in the bottom panels. M1 muscarinic receptor activated by carbachol.
Recent discoveries using our live cell fluorescent sensors:
- T. Buranda, et. al. A High-Throughput Flow Cytometry Screen Identifies Molecules that Inhibit Hantavirus Cell Entry. SLAS Discovery. April 2, 2018.
- H. Zou, M. Schmidt, and R. Gosens PDE8: A Novel Target in Airway Smooth Muscle. American Journal of Respiratory Cell and Molecular Biology. Vol. 58, No. 4. April 01 2018
- T.B. Johnstone, et al. PDE8 is Expressed in Human Airway Smooth Muscle and Selectively Regulates cAMP Signaling by β2AR-AC6. American Journal of Respiratory Cell and Molecular Biology. Dec. 2017
- Tatsuru Togo, Cell membrane disruption stimulates cAMP and Ca2+ signaling to potentiate cell membrane resealing in neighboring cells. Biology Open.
- McCrink KA, Maning J, Vu A, Jafferjee M, Marrero C, Brill A, Bathgate-Siryk A, Dabul S, Koch WJ, Lymperopoulos A. β-Arrestin2 Improves Post-Myocardial Infarction Heart Failure via Sarco(endo)plasmic Reticulum Ca2+-ATPase-Dependent Positive Inotropy in Cardiomyocytes. Hypertension. 2017 Nov; 70: 972-981. Link to Supplemental Data
- Q. Wang, A. N. Pronin, K. Levay, J. Almaça, A. Fornoni, A. Caicedo, V. Z. Slepak. Regulator of G protein signaling Gβ5-R7 is a crucial activator of muscarinic M3 receptor-stimulated insulin secretion. FASEB J. 2017 Jul 7, fj.201700197RR.
- P. Tewson, S.Martinka, C. Berlot, T. Hughes, A.M. Quinn, S. Tillo, Measuring dynamic and real-time cAMP levels using cADDis, a live-cell indicator for Gs and Gi signaling. SLAS 2017.
- SLAS Conference Abstract, 2017. Direct Measurements of Gi signaling and its dynamic interplay with Gs using cADDis, a live-cell assay for cAMP with improved sensitivity.
P. Tewson, A.M. Quinn, T. Hughes. Dynamic Measurements of Gi Signaling in Living Cells. FASEB J. 2016 April, vol. 30 no. 1. Supplement lb478
- J. Almaça, J. Molina, D. Menegaz, A.N. Pronin, A. Tamayo, V. Slepak, P. Berggren, A. Caicedo. Human beta cells produce and release serotonin to inhibit glucagon secretion from alpha cells. Cell Reports, Volume 17, Issue 12, p3281–3291, 2016.
- GLP-1 stimulates insulin secretion by PKC-dependent TRPM4 and TRPM5 activation. J Clin Invest. 2015; 125(12):4714-4728.
- Does PKC activation increase the homologous desensitization of µ opioid receptors? Br J Pharmacy. 2015; 172(2):583-92.
- PKC-dependent Phosphorylation of the H1 Histamine Receptor Modulates TRPC6 Activity. Cells. 2014. 3(2): 247-257.
- New DAG and cAMP Sensors Optimized for Live-Cell Assays in Automated Laboratories. J Biomol Screen. 2015. DOI: 10.1177/1087057115618608
- A multiplexed fluorescent assay for independent second-messenger systems: decoding GPCR activation in living cells. J Biomol Screen. 2013. 18, 797–806Simultaneous Detection of Ca2+ and Diacylglycerol Signaling in Living Cells. PLoS ONE. 2012. DOI: 10.1371/journal.pone.0042791
- Cilia Have High cAMP Levels That Are Inhibited by Sonic Hedgehog Regulated Calcium Dynamics. PNAS 2016. In Press
Fluorescent Assays for Live Cell Discovery