FIG. 2.

(a) Schematic showing that volumetric fraction of erythrocytes (hematocrit or Ht) significantly varies through the entire vascular network, decreasing from systemic blood vessels down to capillaries. (b) Experiments with whole and diluted blood show that the temperature dependent optoacoustic response is scaled proportionally to hematocrit. (c) However, when normalized at 37 °C, it becomes invariant as the curves representing whole and diluted blood coincide. Curves labeled Ht 40 represent whole porcine blood. Curves with reduced Ht indicate that the blood sample was diluted, for example, Ht 20 represents a sample with dilution fraction of ½. Dashed-dotted line marks zero optoacoustic response. Yellow dashed line on the panel (c) shows the universal blood calibration curve $\overline{\mathit{O}\mathit{A}}=-\frac{4\Delta T_{\mathit{m}\mathit{a}\mathit{x}}}{{\left(T_1-T_0\right)}^3}\left(T-T_0\right)\left(T-T_1\right)+\frac{T-T_0}{T_1-T_0}$ with parameters: T₁ = 37 °C—fixed normalization temperature, where normalized optoacoustic intensity $\overline{\mathit{O}\mathit{A}}=1$; T₀ = −12.8 ± 0.5 °C—temperature of zero optoacoustic response estimated directly from the image (N = 8 animal samples); and ΔT_max = 10.8 ± 0.2 °C—maximum nonlinear temperature deviation in the temperature range [T₀ T₁]. If ΔT_max = 0, the calibration is linear. For comparison, normalized Grüneisen parameter of water has maximum nonlinear temperature deviation ΔT_max = 1.9 °C in the range of 4–37 °C. For more details on ΔT_max, see supplementary material.⁴⁵