Edits, Characterization Section

Author: Muaaz-F

docs/source/dvopm_characterization/Images/Axial_Thickness_Intensity.jpg

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 .. _dvopmcharacterization-home:
 
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 Altair DV-OPM Characterization
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+To characterize the performance of the Altair DV-OPM, we evaluated the performance of the illumination train by imaging the sheet profile, and we also imged 1um fluorescent beads to quantify the resolution of the system.
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+Beam Characterization
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+To characterize the beam profile of the illumination train, we imaged the sheet profile in air at a range of axial positions.
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+To characterize the illumination beam, we imaged the light sheet in air while translating the camera through a series of axial positions along the propagation direction, $Y'$. We then selected a fixed lateral region near the center of the sheet, so that the thickness measurement was made from the same representative part of the sheet at every axial position.
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+For each axial plane, the background-corrected intensity profile along the sheet thickness direction, $Z'$, was extracted and fit with a Gaussian function. The fitted Gaussian standard deviation, $\sigma$, was converted to FWHM using $\mathrm{FWHM}=2.35\sigma$ after applying the camera pixel size. The FWHM was subsequently converted to the Gaussian beam waist, $w$, using $w=\mathrm{FWHM}/\sqrt{2\ln2}$, where $w$ corresponds to the $1/e^2$ intensity radius. The best-focus plane was defined as the axial position with the minimum FWHM. From this plane, we measured an FWHM of $17.1~\mu\mathrm{m}$, corresponding to $w_0=14.53~\mu\mathrm{m}$.
+We directly measured the usable axial distance from the experimental waist curve as the range over which $w(Y') \leq \sqrt{2}w_0$, giving a usable distance of approximately $2.00~\mathrm{mm}$.
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+.. figure:: Images/Axial_Thickness_Intensity.jpg
+    :align: center
+    :alt: Sheet profile characterization.
+
+    **Figure 1:** Characterization of the light sheet profile in air along the axial direction.
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+At the best-focus plane, we also characterized the spatial distribution of the sheet. A representative image was used to show the light sheet morphology and its long lateral extent along $X'$. The normalized background-corrected intensity profile along $Z'$ reports the measured sheet thickness, while the lateral profile along $X'$ was calculated by summing the background-corrected intensity along $Z'$ at each lateral position. This provided a measure of the illumination distribution across the approximately $6.5~\mathrm{mm}$ lateral extent of the sheet.
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+.. figure:: Images/Focal_Plane.jpg
+    :align: center
+    :alt: Sheet profile characterization at focal plane.
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+    **Figure 2:** Characterization of the light sheet profile in air at the best-focus plane. A. Camera Image of the light sheet at the best-focus plane. B. Normalized intensity profile along the sheet thickness direction, $Z'$. C. Normalized intensity profile along the lateral direction, $X'$.
 
 Beads
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+To quantify the resolution of the system, we imaged 1um fluorescent beads embedded in agarose.
 
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+We imaged the beads at different axial positions along the propagation direction, $Y'$, and we measured the FWHM of the bead images along the lateral and axial directions to quantify the resolution of the system.
 
-Cells
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+To prepare fluorescent bead samples, we diluted 1 µm YG nanosphere at a concentration of 1:1000 with deionized water and sonicated for 3 minutes to minimize aggregation. The resulting solution was later diluted at a concentration of 1:100 with 2% low-melting agarose for volumetric imaging.
 
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+.. figure:: Images/Beads_All.jpg
+    :align: center
+    :alt: Bead characterization.
 
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+    **Figure 3:** Image of 1um fluorescent beads embedded in agarose, imaged with the Altair DV-OPM. The beads are visible as bright spots against the dark background, and their size and shape can be used to quantify the resolution of the system.
 
-Liver
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+.. figure:: Images/Beads_Single.jpg
+    :align: center
+    :alt: Bead characterization profiles.
 
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+    **Figure 4:** Single Zoomed-in Bead Image in XY and XZ planes.
 
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+We measured the full width at half-maximum (FWHM) values measured from images of 1 µm fluorescent beads (n = 302 beads), binned into 8 sections by lateral position across the field of view and the median values in each section are reported (in micrometers) for the X′, Y′ and Z′ directions. 
 
-Brain
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+.. figure:: Images/Median_FWHM_final.svg
+    :align: center
+    :alt: Bead characterization profiles.
 
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+    **Figure 5:** Median FWHM values measured from images of 1 µm fluorescent beads, binned by lateral position across the field of view.
 
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