Table 3.

Biophysical characterization of GCaMPs

	ϵ^{^a} (M⁻¹cm⁻¹)		QY^{^b}		τ^{^c} (ns)				ΔF/F_o^d pH 7.25	QY*σ₂^{^e} pH 9.5 (+ Ca²⁺)	Peak brightness^{^f} (kcpsm) pH 9.5 (+Ca²⁺)
	pH 7.25 (+Ca²⁺)	pH 9.5 (+Ca²⁺)	pH 7.25 (+Ca²⁺)	pH 9.5 (+Ca²⁺)	pH 7.25 (+Ca²⁺)	pH 7.25 (−Ca²⁺)	pH 9.5 (+Ca²⁺)	pH 9.5 (−Ca²⁺)	ΔF/F_o^d pH 7.25	QY*σ₂^{^e} pH 9.5 (+ Ca²⁺)	Peak brightness^{^f} (kcpsm) pH 9.5 (+Ca²⁺)
EGFP	56,000	57,800	0.73	0.70	2.6	2.6	2.6	2.6		1	13.4
GCaMP2	38,100	54,800 (60,600)	0.53	0.55	2.0	2.1	2.0	2.1	5.7 ± 1.4	0.80	6.7
GCaMP3	37,700	56,300 (66,500)	0.65	0.67	2.8	2.7	2.7	2.6	10.6 ± 1.2	0.83	9.5
GCaMP5A	56,100	58,100 (60,900)	0.65	0.62	2.7	2.8	2.6	2.7	17 ± 4.2	0.89	9.9
GCaMP5D	25,300	48,100 (58,300)	0.67	0.63	2.7	2.7	2.7	2.8	18 ± 3.7	0.78	9.0
GCaMP5G	49,300	58,500 (61,300)	0.67	0.67	2.7	2.8	2.7	2.7	31 ± 9.4	0.96	9.3

↵^aPeak extinction coefficient at 497 nm (485 nm for GCaMP2, 487 nm for EGFP), based on concentration determined by alkali denaturation: 0.1 m NaOH, ϵ = 44,000 m⁻¹cm⁻¹ at 447 nm. Values in parentheses are based on concentration measurements obtained by two-photon FCS at pH 9.5.
↵^bQuantum yield (QY) using fluorescein (QY = 0.93) in pH 11/H₂O as a standard.
↵^cFluorescence lifetime under two-photon excitation (960 nm, 2 mW), fit to a single-exponential decay.
^dCalcium-dependent contrast ΔF/F in pH 7.25 buffer, for 940 nm excitation; SD calculated for eight or more measurements.
↵^eTwo-photon action cross section (QY × two-photon cross section σ₂ per molecule) at 940 nm excitation, relative to EGFP, in pH 9.5 buffer containing 1 mm CaCl₂.
↵^fPeak brightness per emitting fluorophore, in kilocounts per second per molecule (kcpsm), defined as the maximum fluorescence rate divided by the number of emitting fluorophores in the excitation beam volume as determined from FCS measurements, as the laser power is scanned from 1–80 mW at an excitation wavelength of 940 nm.