Practical Aspects of Measuring Intracellular Calcium Signals with Fluorescent Indicators - Methods in Cell Biology - page 113

Table I
Properties of common fluorescent Ca 2 þ indicators a

Absorption maxima
(nm) Emission maxima (nm)

Ca 2 þ -free Ca 2 þ -bound Ca 2 þ -free Ca 2 þ -bound

Indicator type K d (nM)

Nonratiometric
Monomeric
Quin2

115 b

352 332 492 498

Fluo-2/Fluo-8 c

380 - 492 - 514

Fluo-3 400 503 506 526 526

Fluo-4 345 491 494 516 516

Calcium Green-1 TM

190 506 506 531 531

Calcium Green-2 TM

550 506 503 536 536

Calcium Green-5N TM 14 d

506 506 532 532

Oregon Green 488 BAPTA-1 TM

170 494 494 523 523

Oregon Green 488 BAPTA-2 TM

580 494 494 523 523

1.0 d

Rhod-2 556 553 576 576

Calcium Orange TM

328 549 549 575 576

Calcium Crimson TM

185 589 589 615 615

Dextran-conjugated e
Fluo-4 dextran (MW 10,000)

600 494 494 518 518

Calcium Green-1 dextran (MW 3000-70,000) f 240-540 f

509 509 534 534

Oregon Green 488 BAPTA-1 dextran (MW 10,000) 265 496 496 524 524

Ratiometric
Monomeric
Fura-2 224 b

362 335 512 505

Fura Red TM

140 473 436 670 655

250 b

Indo-1 349 331 485 410

Dextran-conjugated e
Fura(-2) dextran (MW 10,000)

240 364 338 501 494

a Data from Tsien (1980), Grynkiewicz et al.

(1985), Minta et al. (1989), Haugland (1992) , and Molecular Probes, The

Handbook (web publication, Invitrogen Corporation).
b E V ective K d in the presence of 1 mMMg 2 þ . (Generally, competition by Mg 2 þ slightly reduces the a Y nity of any indicator
for Ca 2 þ .)
c Di V erent names for the same molecule.
d

m M.
e The K d and absorption and emission maxima of dextran-conjugated indicators can vary from lot-to-lot and is dependent
on the molecular weight of the dextran used as well.
f

K d is reported to be di V erent between low- and high-MW versions: 540 nM for MW 3000, 240-250 nM for MW
10,000 and 70,000.

peaks remain essentially unchanged. Excitation spectra of Fluo-3 ( Minta et al.,
1989 ), a nonratiometric indicator, at saturating and ''zero'' [Ca 2 þ ] are shown in
Fig. 2 . It can be seen that peak excitation occurs at 505 nm irrespective of

whether the indicator is Ca 2 þ -free or Ca 2 þ -bound—the defining characteristic of
a nonratiometric indicator. In contrast, ratiometric indicators exhibit not only
intensity changes with changing [Ca 2 þ ] but the Ca 2 þ -free and Ca 2 þ -bound forms