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Improvements in live cell analysis of G protein coupled receptors using second generation BD calcium assay kits.

Li X, Llorente I, Brasch M - Curr Chem Genomics (2008)

Bottom Line: The kits use a proprietary formulation including a non-fluorescent calcium indicator that becomes activated inside the cell and shows increased fluorescence upon calcium binding.We have compared the next generation BD calcium assay kit product family to previous versions of the formulation, and to other commercially available homogeneous calcium assay kits.The improvements have enabled better performance on the cell lines and receptors that we have tested in all plate formats including 1536.

View Article: PubMed Central - PubMed

Affiliation: Bioimaging Systems, BD Biosciences, Rockville, Maryland, USA. xiao_li@bd.com

ABSTRACT
BD Calcium Assay Kits are designed for cell-based calcium mobilization high-throughput screening assays. The kits use a proprietary formulation including a non-fluorescent calcium indicator that becomes activated inside the cell and shows increased fluorescence upon calcium binding. The formulation includes a signal-enhancing reagent to maximize the signal over background in a homogeneous, no-wash assay format, based on a technology developed at BD. We have compared the next generation BD calcium assay kit product family to previous versions of the formulation, and to other commercially available homogeneous calcium assay kits. The improvements have enabled better performance on the cell lines and receptors that we have tested in all plate formats including 1536.

No MeSH data available.


Dose response curves of pirenzepine in CHO-M1 cells in the presence of 40 nM carbachol at 384-well format. Signal to background are 4.8 and 5.4, IC50 values are 224 nM and 200 nM using the first and second generation kits, respectively.○ BD™ High Performance Calcium Assay Kit (second generation).□ BD™ Calcium Assay Kit (first generation).
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Figure 3: Dose response curves of pirenzepine in CHO-M1 cells in the presence of 40 nM carbachol at 384-well format. Signal to background are 4.8 and 5.4, IC50 values are 224 nM and 200 nM using the first and second generation kits, respectively.○ BD™ High Performance Calcium Assay Kit (second generation).□ BD™ Calcium Assay Kit (first generation).

Mentions: Antagonsit assay was also conducted using both generations of calcium assay kits. Pirenzepine, a known muscarinic receptor inhibitor, was incubated with CHO cells overexpressing rat muscarinic receptor M1 for 5 minutes before the cells were applied with carbachol at EC80 value (40 nM in this case). As shown in Fig. (3), the assay using the second generation calcium assay kit delivered higher signal to background than the assay using the first generation kit. The IC 50 value remained similar between the two assays.


Improvements in live cell analysis of G protein coupled receptors using second generation BD calcium assay kits.

Li X, Llorente I, Brasch M - Curr Chem Genomics (2008)

Dose response curves of pirenzepine in CHO-M1 cells in the presence of 40 nM carbachol at 384-well format. Signal to background are 4.8 and 5.4, IC50 values are 224 nM and 200 nM using the first and second generation kits, respectively.○ BD™ High Performance Calcium Assay Kit (second generation).□ BD™ Calcium Assay Kit (first generation).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2803433&req=5

Figure 3: Dose response curves of pirenzepine in CHO-M1 cells in the presence of 40 nM carbachol at 384-well format. Signal to background are 4.8 and 5.4, IC50 values are 224 nM and 200 nM using the first and second generation kits, respectively.○ BD™ High Performance Calcium Assay Kit (second generation).□ BD™ Calcium Assay Kit (first generation).
Mentions: Antagonsit assay was also conducted using both generations of calcium assay kits. Pirenzepine, a known muscarinic receptor inhibitor, was incubated with CHO cells overexpressing rat muscarinic receptor M1 for 5 minutes before the cells were applied with carbachol at EC80 value (40 nM in this case). As shown in Fig. (3), the assay using the second generation calcium assay kit delivered higher signal to background than the assay using the first generation kit. The IC 50 value remained similar between the two assays.

Bottom Line: The kits use a proprietary formulation including a non-fluorescent calcium indicator that becomes activated inside the cell and shows increased fluorescence upon calcium binding.We have compared the next generation BD calcium assay kit product family to previous versions of the formulation, and to other commercially available homogeneous calcium assay kits.The improvements have enabled better performance on the cell lines and receptors that we have tested in all plate formats including 1536.

View Article: PubMed Central - PubMed

Affiliation: Bioimaging Systems, BD Biosciences, Rockville, Maryland, USA. xiao_li@bd.com

ABSTRACT
BD Calcium Assay Kits are designed for cell-based calcium mobilization high-throughput screening assays. The kits use a proprietary formulation including a non-fluorescent calcium indicator that becomes activated inside the cell and shows increased fluorescence upon calcium binding. The formulation includes a signal-enhancing reagent to maximize the signal over background in a homogeneous, no-wash assay format, based on a technology developed at BD. We have compared the next generation BD calcium assay kit product family to previous versions of the formulation, and to other commercially available homogeneous calcium assay kits. The improvements have enabled better performance on the cell lines and receptors that we have tested in all plate formats including 1536.

No MeSH data available.