ASAP2s

Higher-sensitivity ASAP1 variant that trades speed for larger fluorescence responses.

Category: VSD-cpGFP GEVI
Published: 2017-07-27 — eLife 2017
Derived from: ASAP1
Response polarity: negative-going (fluorescence decreases on depolarization)
Spectrum: excitation peak 490 nm, emission peak 511 nm (ASAP2s)
Two-photon compatible: yes
Plasmid: Addgene #101274
Tags: cpGFP, Two-photon

Headline values (as shown in the GEVIBench table)

Metric	Value	Definition
Brightness (B/B_EGFP)	0.43× EGFP	Molecular brightness relative to EGFP (extinction coefficient × quantum yield, or a measured ratio). Sensors not measured against EGFP directly are placed through a chain of pairwise brightness comparisons, combined by geometric mean.
Kinetics τ_on / τ_off (ms)	7.8 / 14 ms	Activation / decay time constants in ms (amplitude-weighted mean τ for multi-exponential fits). The value shown averages the measurements made at physiological temperature (≥33 °C); if there are none, it is the measurement closest to 33 °C.
ΔF/F per 100 mV	-38.7% (median of 4 measurements: −36%)	Steady-state fluorescence change for a 100 mV depolarization step (−70 → +30 mV); the sign gives the polarity. The table shows the first listed measurement; the median across measurements (one-photon preferred) is used for sorting.
ΔF/F per mV	0.15%	Subthreshold sensitivity: fluorescence change per mV near rest (−90 to −50 mV). Where no value is reported directly, it is estimated from the F–V curve.
ΔF/F per AP	12.2% (median of 3 measurements: 9.6%)	Fluorescence change per single action potential in neurons. The table shows the first listed measurement; the median across measurements (one-photon preferred) is used for sorting.
AP width (FWHM, ms)	15 ms	Full width at half maximum of the optical spike waveform.
Photostability t75% @ 100 mW/mm²	5.5 s	Time for fluorescence to fall to 75% of its initial value under one-photon illumination, linearly dose-scaled to 100 mW/mm² (t50% is used where the 75% crossing falls inside a fast initial transient). Two-photon curves are not dose-scaled and are excluded.
Papers using it (N_used)	13	Number of published research papers, tracked by GEVIBench, that used the sensor for an experiment (a popularity / track-record measure).

Measurements and sources

Kinetics

τ_on 30.6 ms, τ_off 65.8 ms at 22°C — Chamberland 2017, Table 1
τ_on 3.58 ms, τ_off 6.99 ms at 32-35°C — Villette 2019, Table 1
τ_on 11.99 ms, τ_off 21.07 ms at 32-35°C — Liu 2022, Table S2
τ_on 16.71 ms, τ_off 29.51 ms at 21-23°C — Liu 2022, Table S2

ΔF/F per 100 mV

-38.7%, 1P — Chamberland 2017, Fig. 1C
-36%, 1P — Villette 2019, Table 1
-28.5%, 1P — Lu 2023, Fig. 2c
-33.3%, 2P — Liu 2022, Fig. 2C

ΔF/F per mV (subthreshold)

0.15%/mV, 2P (Subthreshold voltage sensitivity ≈ 0.15 %/mV, from the Fig. 5G peak-response-vs-depolarization plot in organotypic hippocampal slices under 2P (462 Hz). ASAP2s ΔF/F peaks were ~0.9%/1.35%/2.2%/3.15% for 5/10/15/20 mV subthreshold depolarization waveforms (least-squares slope 0.154 %/mV, read from the bar graph). This is a 2P slice measurement of responses to subthreshold-waveform commands (not a 1P near-rest voltage-clamp ramp), so it may not be directly comparable to 1P subthreshold slopes.) — Chamberland 2017, Fig. 5G

ΔF/F per action potential

AP width (FWHM)

21.1 ms at 3700 Hz sampling, Organotypic hippocampal slice cultures, current-triggered single AP, two-photon RAMP (FEVIR), 2P, 22°C — Chamberland 2017, Fig. 6E
17.4 ms at 1000 Hz sampling, Rat, cultured hippocampal neurons, in vitro, 1P — Chamberland 2017, Fig. 2J
17.3 ms at undefined Hz sampling, Rat, cultured hippocampal neurons, current-evoked single AP, room temperature, in vitro, 1P, 22°C — Villette 2019, Fig. 2F
12.3 ms at 500 Hz sampling, Mouse, cultured hippocampal neurons, current-triggered single AP, in vitro, 1P — Milosevic 2020, Fig. 1B
6.7 ms at 987 Hz sampling, HEK293A cells, optical response to a single AP-like command waveform (2-ms FWHM, +30 mV peak, −70 mV baseline), whole-cell voltage clamp, in vitro, 1P, 25°C — Lu 2023, Fig. 2f
7.6 ms at 440 Hz sampling, HEK293A cells, optical response to a single AP-like command waveform (2-ms FWHM), whole-cell voltage clamp, in vitro, 2P, 22°C — Liu 2022, Fig. 2F

Brightness comparisons

1.11× the brightness of ASAP3 — Liu 2022, Fig. 2I
1× the brightness of ASAP1 — Chamberland 2017, Fig. 1—figure supplement 2A
0.45454545454545453× the brightness of JEDI-1P — Lu 2023, Fig. 2h
0.6666666666666666× the brightness of JEDI-2P — Liu 2022, Fig. 2I

Photobleaching

curve only (did not reach 75% within the measured window) at 46 mW/mm², 1P — Lu 2023, Fig. S4
t50% = 3 s at 15 mW/mm², 1P — Liu 2022, Fig. S4G
t75% = 50 s at 11 mW/mm², 1P — Chamberland 2017, Fig. 1—figure supplement 3A
t75% = 47.7 s at 24 mW/mm², 1P — Chamberland 2017, Fig. 2—figure supplement 3B
t75% = 20.4 s at 6.4 mW at objective back aperture (2P, 920 nm), 2P — Chamberland 2017, Fig. 1—figure supplement 4A

Fluorescence–voltage (F–V) curve

ΔF/F (%) at each membrane potential, normalized to 0 at −70 mV: -100 mV: 18%; -90 mV: 13%; -80 mV: 6%; -70 mV: 0%; -60 mV: -6%; -50 mV: -11%; -40 mV: -17%; -30 mV: -21%; -20 mV: -25%; -10 mV: -28%; 0 mV: -32%; 10 mV: -34%; 20 mV: -37%; 30 mV: -39%; 40 mV: -40%; 50 mV: -42% — Chamberland 2017, Fig. 1C

Research papers using ASAP2s (13)

Fast two-photon imaging of subcellular voltage dynamics in neuronal tissue with genetically encoded indicators — Chamberland S, Yang HH, et al., eLife 2017 (HEK293A cells; rat hippocampal neurons; Drosophila L2 neurons; mouse hippocampal slice cultures, in vitro/in vivo)
In Vitro Testing of Voltage Indicators: Archon1, ArcLightD, ASAP1, ASAP2s, ASAP3b, Bongwoori-Pos6, BeRST1, FlicR1, and Chi-VSFP-Butterfly — Milosevic MM, et al., eNeuro 2020 (Mouse, cortex; HEK293 cells, in vitro)
Ultrafast two-photon imaging of a high-gain voltage indicator in awake behaving mice — Villette V, et al., Cell 2019 (Rat, hippocampal neurons; HEK293A cells; mouse, cerebellar slices, in vitro)
Imaging of Evoked Cortical Depolarizations Using Either ASAP2s, or chi-VSFP, or Di-4-Anepps, or Autofluorescence Optical Signals — Milicevic KD, et al., Journal of Integrative Neuroscience 2023 (Mouse, cortex; ASAP2s-transgenic brain slices, ex vivo)
CaMKII Measures the Passage of Time to Coordinate Behavior and Motivational State — Thornquist SC, et al., Neuron 2019 (Drosophila, abdominal ganglion neurons, ex vivo)
Biochemical evidence accumulates across neurons to drive a network-level eruption — Thornquist SC, et al., Molecular Cell 2021 (Drosophila, Corazonin neurons, ex vivo)
Drosophila TRPγ is required in neuroendocrine cells for post-ingestive food selection — Dhakal S, et al., eLife 2022 (Drosophila, DH44 neuroendocrine cells, in vivo)
Triclosan disrupts immune cell function by depressing Ca2+ influx following acidification of the cytoplasm — Sangroula S, et al., Toxicology and Applied Pharmacology 2020 (RBL mast cells, in vitro)
Real-time detection of changes in yeast plasma membrane potential using genetically encoded voltage indicator proteins — Limapichat W, et al., FEMS Yeast Research 2020 (Saccharomyces cerevisiae, in vitro)
A dark quencher genetically encodable voltage indicator (dqGEVI) exhibits high fidelity and speed — Alich TC, et al., Proceedings of the National Academy of Sciences 2021 (HEK293T cells; rat hippocampal neurons, in vitro)
Sustained deep-tissue voltage recording using a fast indicator evolved for two-photon microscopy — Liu Z, Lu X, et al., Cell 2022 (HEK293A cells; mouse hippocampal neurons, in vitro)
Widefield imaging of rapid pan-cortical voltage dynamics with an indicator evolved for one-photon microscopy — Lu X, Wang Y, Liu Z, et al., Nature Communications 2023 (HEK293A cells; HEK293-Kir2.1 cells; rat cortical and hippocampal neurons, in vitro)
Physiological features of parvalbumin-expressing GABAergic interneurons contributing to high-frequency oscillations in the cerebral cortex — Milicevic KD, et al., Current Research in Neurobiology 2023 (Mouse, cortex; PV-ASAP2s transgenic brain slices, ex vivo)

Data and citation

llms.txt — summary for language models
llms-full.txt — every sensor in full (Markdown)
gevis.json — all values and measurements with sources (JSON)
gevis.csv — one row per sensor (CSV)

Cite GEVIBench as: Adam E. Cohen and Daozheng Gong (2026). Voltage imaging: A practical guide. Neuron. https://doi.org/10.1016/j.neuron.2026.08.002

Generated from the GEVIBench database on 2026-10-12.