Figure 2.

Histograms of fluorescence intensity of spots and difference intensity observed by TIRFM. (a) Histograms of maximum intensity in asolectin liposomes at pH 6 (n = 1850) fitted with three Gaussian functions shown in Eq. 1 (μ = 154, σ = 45, A₁ = 186, A₂ = 59, A₃ = 6). (b–d) Histograms of the fluorescence decrease upon photobleaching of Alexa594 in F_high (b), F_low (c), and unidentified state because of photobleaching before the fluorescence alternation (d). They were fitted with the single Gaussian functions [n = 676, μ = −160, σ = 51, A = 99 for (b), n = 471, μ = −126, σ = 46, A = 78 for (c), and n = 703, μ = −145, σ = 38, A = 134 for (d)]. (e–g) Difference intensity histograms for photoactivated rhodopsin (n = 924) (e), denatured rhodopsin (n = 505) (f), and free Alexa594 molecules in solution (n = 129) (g). The difference intensity histogram in (e) was well fitted with two mirror-imaged pairs of Gaussian functions (Eq. 2: μ₁ = ±0.304, σ₁ = 0.111, A₁ = 0.014, and μ₂ = ±0.167, σ₂ = 0.049, A₂ = 0.037), whereas those in (f) and (g) were fitted with a single mirror-imaged pair of Gaussian functions [μ = ±0.344, σ = 0.112, A = 0.011 (f), and μ = ±0.377, σ = 0.127, A = 0.011 (g)].