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Camp glo kit

Manufactured by Promega
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The CAMP-Glo kit is a laboratory tool designed to detect and quantify cyclic AMP (cAMP) levels in biological samples. It utilizes a bioluminescent assay to measure cAMP, which plays a crucial role in various cellular signaling pathways.

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Lab products found in correlation

Forskolin, by Santa Cruz Biotechnology (1 mentions) Synergy neo, by Agilent Technologies (1 mentions) Camp glo assay, by Promega (1 mentions) Flexstation 3, by Molecular Devices (1 mentions) Forskolin, by Merck Group (1 mentions) 96 well plate, by Thermo Fisher Scientific (1 mentions) Infinite f200 pro illuminometer, by Tecan (1 mentions)

13 protocols using camp glo kit

1

Cyclic AMP Measurement in GPER Agonist Assay

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Suit2-007 cells were seeded in a 96-well plate. Then, 24 h after seeding, they were incubated with 100 μM IBMX (3-Isobutyl-1-methylxanthine; Sigma-Aldrich) for 20 min, followed by incubation with 10 μM forskolin (sc-3562; Santa Cruz Biotechnology) and 100 μM IBMX or 10 μM forskolin, 100 μM IBMX and 1 µM G1 GPER agonist for a further 20 min. Cyclic AMP levels were measured using cAMP-Glo kit (V1501, Promega, Madison, WI, USA) based on an inversely proportional bioluminescent luciferase reaction. Cyclic AMP levels are represented as inversed luminescence values normalised to control conditions (IBMX + forskolin). forskolin was used to increase the baseline cAMP levels past the assay threshold and to potentiate the receptor to more accurately determine the response to the agonist as described in [37 (link)].
Rice A., Cortes E., Lachowski D., Oertle P., Matellan C., Thorpe S.D., Ghose R., Wang H., Lee D.A., Plodinec M, & del Río Hernández A.E. (2020). GPER Activation Inhibits Cancer Cell Mechanotransduction and Basement Membrane Invasion via RhoA. Cancers, 12(2), 289.
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2

Measuring cAMP Levels in HaCaT Cells

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HaCaT cells were seeded in 96-well plates at a density of 5×103 cells/well. After 24 hours, the cells were stimulated for 10 minutes with different concentrations (10, 25, and 50 µM) of α-ionone, of forskolin (10 µM) or solvent (0.1% DMSO) only. Forskolin was used as a positive control and the solvents served as the vehicle control. The cAMP levels in stimulated cells were determined using the cAMP-Glo™ Kit (Promega, Madison, WI, USA) according to the manufacturer’s instructions and measured via a plate reader (Packard; PerkinElmer, Waltham, MA, USA).
Yang D., Fu M., Zhao Q., Wang Y., Li T., Feng B., Li E., Nishijima Y., Sun Z, & Hu Z. (2023). α-ionone promotes keratinocyte functions and accelerates epidermal barrier recovery. Annals of Translational Medicine, 11(8), 297.
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3

Evaluating cAMP Levels in 3T3-L1 Cells

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3T3-L1 (2X104 cells/well) were plated in white clear bottom 96 well plates (BD 353377) and grown overnight. Cells were serum starved for 3 h. Cells were transferred to 100 μL KRB buffer with 4% fatty acid free BSA and pre-treated with 0.5 mM IBMX for 10 min followed by 15 min and 100 min of IBMX± CRACE (500 μg/mL equivalent) treatment. Intracellular cAMP was measured using cAMP-Glo Kit (Promega V1501).
Borah A.K., Singh A., Yasmin R., Doley R., Mattaparthi V.S, & Saha S. (2019). 1α, 25-dihydroxy Vitamin D3 containing fractions of Catharanthus roseus leaf aqueous extract inhibit preadipocyte differentiation and induce lipolysis in 3T3-L1 cells. BMC Complementary and Alternative Medicine, 19, 338.
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4

Transfection and FACS Sorting of CHO Cells

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Chinese hamster ovary (CHO) cells (2.5 × 106) were seeded on a 10 mm diameter dish. After 24 h cells were transfected according to the manufacturer's instructions (FuGENE HD; cat. no. 0470969100; Roche Diagnostics GmbH, Mannheim, Germany) with (1) pmaxGFP (kindly provided by Paul Heppenstall, EMBL), (2) pcDNA3-Htr1a-P2A-tdTomato, or (3) a mixture of pmaxGFP and pcDNA3-Htr1a. Forty 8 h later cells were harvested by trypsinization, resuspended in single cell suspension and sorted by FACS using GFP (1 and 3) or tdTomato (2) channels. Fifteen thousand cells were seeded per well in a 96-well plate. Eighteen hours later cAMP assay was performed using cAMP-Glo kit according to manufacturer instruction (Promega Madison, USA).
Piszczek L., Piszczek A., Kuczmanska J., Audero E, & Gross C.T. (2015). Modulation of anxiety by cortical serotonin 1A receptors. Frontiers in Behavioral Neuroscience, 9, 48.
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5

PGE2-Induced cAMP Measurement in hBMSCs

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hBMSCs were seeded at 15,000 cells/cm2 in 96-well plates, and induced to undergo osteogenesis for 3 days. Medium was then replaced with PBS containing PGE2 (100 nM) for up to 15 min. Measurement of intracellular cAMP levels in hBMSCs was then performed using the cAMP-Glo Kit (Promega, Dübendorf, Switzerland) according to the manufacturer’s protocol. The resulting luminescence was measured using a multiplate reader and cAMP concentration calculated according to manufacturer’s protocol (Tecan, Männedorf, Switzerland).
Mirsaidi A., Tiaden A.N, & Richards P.J. (2017). Prostaglandin E2 inhibits matrix mineralization by human bone marrow stromal cell-derived osteoblasts via Epac-dependent cAMP signaling. Scientific Reports, 7, 2243.
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6

cAMP-Glo Assay for Cell Signaling

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One hundred microlitres of 50 000 cells ml−1 cell suspensions of individual cell lines were added in wells in white-walled 96-well plates (Thermo-Fisher Scientific, catalogue no. 07-200-628) and cultured for 72 h. The wells were washed twice with PBS and then treated with 0.1 nM CT holotoxin at 37°C for 60 min (List Biologicals) in complete induction buffer provided in the cAMP-Glo kit (Promega, catalogue no. V1501). The assay was performed following the manufacturer's protocol (cAMP-Glo assay; Promega). The luminescence values were obtained using a Synergy Neo microplate reader (BioTek, Winooski, VT, USA).
Sethi A., Wands A.M., Mettlen M., Krishnamurthy S., Wu H, & Kohler J.J. (2019). Cell type and receptor identity regulate cholera toxin subunit B (CTB) internalization. Interface Focus, 9(2), 20180076.
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7

Quantifying cAMP Levels in OPCs

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To assess cAMP levels in Ift88 cKO OPCs and controls, we used the cAMP-Glo kit (Promega). OPCs were seeded into 12-well plates at a density of 50,000 cells per well in media containing PDGF-AA as described above. PDGF-AA was then removed from media for 4 h and incubated in cAMP-Glo Lysis Buffer and transferred to 96 well plates. cAMP-Glo assay was performed according to manufacturer’s instructions, and plates were read using a microplate reader luminometer.
Bönnemann C.G., Cox G.F., Shapiro F., Wu J.J., Feener C.A., Thompson T.G., Anthony D.C., Eyre D.R., Darras B.T, & Kunkel L.M. (2000). A mutation in the alpha 3 chain of type IX collagen causes autosomal dominant multiple epiphyseal dysplasia with mild myopathy. Proceedings of the National Academy of Sciences of the United States of America, 97(3), 1212-1217.
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8

ARPE19 cAMP Accumulation Assay

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The ARPE19 cells were plated in two 96-well plates at a density of 50,000 cells/ well in 85 μl of DMEM medium containing 10% FBS and antibiotics. Sixteen hours after seeding the cells were treated with SNAP-37889 at 10 μM concentrations for 30 min. Next, galnon at 0.001, 0.1, or 1 μM was added for 30 min. Levels of accumulated cAMP were detected with the cAMP-Glo™ kit (Promega) following the manufacturer’s protocol. The luminescence signal was recorded with a FlexStation 3 plate reader (Molecular Devices). The cAMP provided by the kit was used to prepare the standard curve. The percentage of cAMP was calculated for each condition, assuming the cAMP level detected in the non-treated cells as 100%. Each condition was performed in triplicate and the experiment was repeated.
Ortega J.T., Parmar T, & Jastrzebska B. (2023). Galanin receptor 3 - a new pharmacological target in retina degeneration. Pharmacological research, 188, 106675.
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9

cAMP Activity Assay for Differentiated EBs

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Day 4 EBs were dissociated and replated in 96-well plates. The next day, differentiated cells were treated with activators, inhibitors, or DMSO as control. Cells were harvested after 24-hr treatment. The levels of cAMP activity were determined using the cAMP-Glo Kit (Promega, V1501), according to the manufacturer’s instruction, and using the EnVision automated microplate reader system (Perkin Elmer).
Tsai S.Y., Maass K., Lu J., Fishman G.I., Chen S, & Evans T. (2015). Efficient Generation of Cardiac Purkinje Cells from ESCs by Activating cAMP Signaling. Stem Cell Reports, 4(6), 1089-1102.
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10

Odorant-Induced cAMP Measurement

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The cells were seeded into 96-well plates (ThermoFisher Scientific) and cultivated for 48 h. After cultivation, the cells were stimulated for 30 min with the odorants diluted to different concentrations (100, 250, 500, and 1,000 μM). Forskolin (10 μM, Sigma-Aldrich) was used as a positive control and the solvents served as the negative control. The cAMP levels were determined using the cAMP-Glo™ Kit (Promega) according to the manufacturer’s protocol and measured via a plate reader (Packard, PerkinElmer, Waltham, United States). The determined values were normalized to the control (0.1% ethanol/DMSO).
Weidinger D., Jovancevic N., Zwanziger D., Theurer S., Hönes J., Führer D, & Hatt H. (2021). Functional Characterization of Olfactory Receptors in the Thyroid Gland. Frontiers in Physiology, 12, 676907.
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