Epi-luminescence imaging of a single UCNP using a “blood-immersion” objective.
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Abstract
<p>(A) A photograph of the hemolyzed blood layer between the objective and cover slip. (B) Absorption spectrum of the hemolyzed blood (red solid curve) and UCNP emission spectrum (green dashed curve). (C) Low-magnification images of the UCNP sample recorded through the eyepiece port using the water- (top) and blood- (bottom) immersion objective. The dried UCNP colloid rims appeared green (top) and red (bottom) due to the green light absorption by blood. (D) Epi-luminescence microscopy image of the UCNP constellation identified in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0063292#pone-0063292-g002" target="_blank">Figure 2C</a>, imaged using the blood-immersion objective. The single UCNP is clearly observable, although blurred. The EMCCD camera settings and excitation parameters were equivalent to these of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0063292#pone-0063292-g002" target="_blank">Figure 2</a>. The pixel values were converted to photons/second (ph/s) and color-coded using the look-up bar in (D).</p