Table 1 Properties of the probes

Dye/dye-protein conjugate	Absorption λ_max (nm)	Emission λ_max (nm)	Apparent pK_a	Q.E. (Φ_fl) of 2-Me analog
2-HM OR	554	572	9.58_± 0.06	0.35^a
2-HM SeR	586	604	7.86_± 0.10	N.D.
2-HM SiR	650	664	5.72_± 0.08	0.30^a
2-HM GeR	639	653	5.70_± 0.03	0.34^a
2-HM PGeR	646	658	5.44_± 0.05	0.34^a
2-HM IGeR	679	693	6.24_± 0.10	0.25^b
2-Me GeR	636	651
Her-GeR	643	656
Her-HMIGeR	686	700	6.61_± 0.10
Her-CypHer	650	665	6.78_± 0.08
Avi-GeR	643	656
Avi-HMIGeR	686	700	5.46_± 0.08
Avi-CypHer	651	666	6.86_± 0.03

Abs_max, Em_max (in acidic conditions in which absorbance or fluorescence intensity reach maximum values), apparent pK_a (in 100 mM NaPi buffer), and fluorescence quantum efficiency (Φ_fl) of the 2-Me analog of each probe in PBS (pH 7.4) are summarized. For each rhodamine derivative, apparent pK_a values were calculated from the absorbance; see Supplementary Figs. 1 and 2 for values calculated from the fluorescence, which are almost the same. For the probe-Herceptin/avidin conjugates, apparent pK_a values were calculated by fluorescence
^aCresyl violet in MeOH (Φ_fl = 0.54) as a standard
^bCy5.5 in PBS (Φ_fl = 0.23) as a standard.

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