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«Novel Biophotonic Imaging Techniques for Assessing Women’s Reproductive Health by Tyler Kaine Drake Department of Biomedical Engineering Duke ...»

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An opaque flange is then adjusted to eliminate ambient light and the tube is zero- positioned (0° oriented vertically, and 0- mm corresponding to the endoscope fully advanced towards the fornix).

The tube is then locked into place and remains stationary throughout the scan.

The physician then manually advances the endoscope inside the tube distally, capturing mLCI data every 1 mm and fluorimetric data every 5 mm.

This sampling method is used to achieve equal coverage of both modalities in the axial direction as the fields of view differ between modalities.28, 71 Once an axial scan is completed, the endoscope is rotated 45° and the scan continues back towards the fornix as shown in Figure 3.16.

This is continued until the entire 360° mapping of the vagina is completed.

–  –  –

57 After the background scan, 3.5 mL of fluorescein- labeled Replens gel is inserted into the subject’s vagina.

A waiting period then occurs where the subject participates in a specific protocol.

In the study presented here, designed as a feasibility study, the subject is seated for 1 min, stands for 1 min, sits for 1 min, and then is supine for 7 min.

The physician then inserts the probe assembly into the subject’s vagina to the same axial depth as the background scan, and the scan procedure described above is repeated.

Approximately 750 regions are sampled with the mLCI device in total, each region consisting of 4 A- scans for a total of about 3000 A- scans/patient.

The typical length of a procedure averaged 49 min/patient (maximum = 61 min;

minimum = 36 min).

In addition to the data processing outlined in Sections 3.2.3 and 3.2.4, additional steps must be taken to reconcile the fluorimetric and mLCI data since the mLCI data is sampled 5-  times more frequently and offset due to probe design as in Figure 3.11(c).

First, for each fluorimetric point, 5 consecutive mLCI scans located about the center of the fluorimetric coordinate are averaged.

The mLCI dataset is then rotated by 180° and shifted

- 20 mm to compensate for the geometric offset.

Both the mLCI background and test scan are processed in this manner.

A point- by- point subtraction of the background mLCI scan from the test mLCI scan is then performed to remove any effects from the non- labeled gel used during insertion of the probe.

Coating thickness distribution obtained from in vivo data is presented as a topological plot in Figure 3.17, where the colorbar corresponds to coating depth.

The 0° 58 point is again aligned with a vertical orientation, and 0 mm corresponds to the area closest to the fornix.

The blank area in the fluorimetric data, Figure 3.17(a), results from the offset of the modalities, as the mLCI imaging module in Figure 3.11(c) prevents the fluorimetric endoscope from advancing entirely towards the fornix, thus missing the deepest 20 mm of fluorimetric data.

–  –  –

As a summary metric, the fraction of scanned vaginal tissue with coating is found for both modalities.

The fluorimetric system has a value of 0.4453 and the mLCI system has a value of 0.4500 for a difference of 1.06%.

The fraction of scanned vaginal tissue with a coating greater than 50 µ;m is also determined because this thickness is used as the threshold for minimum coating thickness needed for HIV protection11.

The fluorimetric device has a value of 0.2891 and the mLCI instrument yields a value of 0.3265 for an absolute difference of 12.9%.

Contour lines of the fluorimetric distribution can then be

–  –  –

Figure 3.18(a).

The distributions appear to correlate spatially, as the areas of coating overlap within the first 55 mm of gel distribution.

At thicker gel distributions ( 600 µ;m) the processed mLCI data tends to yields lower values, as the instrument has some loss of signal at such depths in the in vivo application.

However, since the investigations here focus on assessing a minimum thickness, this upper threshold does not alter the analysis.

–  –  –

60 Figure 3.18(b) shows an example cross- sectional image constructed from consecutive A- scans for 1 cm of axial position across a tissue.

Unlike OCT, the mLCI device is not constructed to produce such B- scans as it sacrifices data density for broad tissue coverage.

However, physical features can still be seen in the image, as the reflection from the outer surface of the tube is seen near the top, and reflections from the tissue surface also appear.

The thickness of the gel can then be calculated as the difference between the two peaks, divided by the index of the gel.

3.2.4 Dual-modality instrument updates Following initial human in vivo vaginal imaging studies, the clinical optical imaging instrument was updated to improve its performance as applied in the exam room.

The device was modified in the following three

ways:

(1) the distal probe tip was redesigned eliminating the axial offset of the modalities’ field of views, (2) new epoxy outer tubes replaced the polycarbonate tubes, and (3) equipment was consolidated onto a single cart.

These improvements will allow more accurate data collection and reduce the imaging time of the instrument.





The original design of the mLCI imaging optics, as shown in Figure 3.11(c) and Figure 3.13(c), include a module mounted to the distal end of the fluorimetry probe.

While this allows simultaneous imaging, a 20 mm axial and 180 degree azimuthal offset existed between the modalities’ field of views.

Digital correction was done to rectify this issue, but because of probe geometry, mLCI images an additional 20- mm towards the 61 fornix then fluorimetry, as shown in Figure 3.19.

Notice the data dropout in Figure 3.19, where no fluorimetry data in part (a) is captured at the deepest portion of the vagina, from 0 – 20 mm.

In many gel distributions, the majority of the inserted volume is in this critical area during imaging, so we wish to eliminate this offset.

–  –  –

A custom probe tip was designed and built that eliminates the 20 mm axial offset, but retains the 180 degree azimuthal offset.

The design is shown in Figure 3.20, where part (a) shows that the mLCI imaging module is now inserted into a cutout section of the endoscopic tip, instead of mounted to the distal end.

This rectifies the axial offset and protects the mLCI optics.

62

–  –  –

Figure 3.20:

Photograph of the optical imaging instrument distal probe tip.

Part (a) shows a top down view of the mLCI imaging optics.

Part (b) shows a side view of the fluorimetry window while (c) represents a 180- degree azimuthal rotation of (b) revealing the mLCI imaging optics.

The original polycarbonate tube used for the trials had many inhomogeneities, resulting in about 25% of mLCI scans not containing useable data.

It is hypothesized that two problems produce these unreliable scans.

First, when gel thicknesses are larger, greater than approximately 600 µ;m, the mLCI device lacks sufficient imaging depth to measure these thicknesses in the in vivo environment.

Secondly, the polycarbonate tubes were machined and polished for the original fluorimetry study by Henderson et al., published in 2005, and the thickness and optical properties of the tube vary both axially and azimuthally28.

Fluorimetric measurements are less susceptible to small local inhomogeneities in the polycarbonate tubes as the signal is integrated over an approximately 1 cm field of view.

However, mLCI has six measurement points that

–  –  –

micron scale.

Thus, local changes in the optical properties of the polycarbonate tube greatly affect the fidelity of the mLCI signal.

Figure 3.21, below, shows the location of failed mLCI scans obtained from the imaging data in Figure 3.

19(b).

–  –  –

In Figure 3.21(c), many failed scans exist around the 90° angle and 30- 50 mm depth (orange ROI in (c)) which corresponds to the large depths (~1200 µ;m) visible in the fluorimetry data (Figure 3.19(a)).

However, there still are many failed scans scattered seemingly randomly throughout the map due to the polycarbonate tube optical inhomogeneities.

To address this concern, new epoxy- based protective tubes (Epoxy Technology;

Epo- Tek 301) have been created and a complete mLCI scan of a tube (8 angles azimuthally;

100 cm axially) have been performed to test thickness variations.

The result of this scan is presented in Figure 3.22, where the A- scan peak due to reflection from the 64 outer tube surface is prominent between 500- 600 µ;m.

The figure results from 880 A-  scans overlaid on a common x- axis.

–  –  –

The absolute depth of the peak at 500- 600 µ;m will not reduce the depth range of the gel thickness scan, as it can be adjusted by moving the reference arm.

However, the relative change in position from peak to peak reveals variations in the optical thickness of the epoxy tube.

The tube thickness measurements had a standard deviation of 21.77 µ;m, and a range of 89.87 µ;m (max = 583.99;

min = 540.79), which indicates that the epoxy tube is very optically uniform.

Finally, the footprint of the cart was reduced by combining the fluorimetry video system with the mLCI control laptop.

The original device used a VCR to record video at

–  –  –

onto the video.

A CRT monitor was also used to monitor this video in the exam room.

These devices have been removed from the optical imaging instrument and replaced with a video card, which interfaces with the mLCI laptop.

Video and encoder positioning data is then displayed in real time on the laptop.

These changes allowed the entire control system to be consolidated onto a single cart as shown in Figure 3.23(a).

Part (b) of Figure 3.23 shows the endoscopic probe mounted onto its positioning cart, ready for clinical study.

–  –  –

66

3.3 Summary The development of the dual- modality optical imaging instrument was covered in Chapter 3.

The initial LCI method, outlined by Braun et al., was transformed into a 6-  channel mLCI system in order to increase tissue coverage without mechanical scanners typical of OCT systems.26 The optical properties of this invention, such as OSNR, resolution, falloff, and cross- talk, were characterized.

Calibration measurements were discussed which used the test socket to verify the linearity and accuracy of the system.

Phantom measurements further verified the ability of the benchtop mLCI device to accurately measure gel distributions atop a tissue- like surface.

A clinical dual- modality optical imaging instrument was then designed and built by combining an existing fluorimetry device used by Henderson et al.

with the mLCI instrument.27, 28 A compact mLCI imaging module was created in order to allow simultaneous imaging by the modalities.

The dual- modality device’s optical properties were first verified, followed by calibration socket measurements.

Initial in vivo validation was provided in Section 3.2.3.

Finally, several improvements were made on the dual- modality instrument which increased both measurement fidelity and ease of operation in the clinical setting.

These changes are discussed in the last section of the chapter, Section 3.2.4.

67 4 In vivo dual-modality optical imaging instrument clinical study

4.1 Introduction

Chapter

3 outlined the details of the instrumentation of a dual- modality optical imaging device, which obtains simultaneous measurements from fluorimetry and mLCI.

The progression of the dual- modality optical imaging instrument was described from a single channel device to a benchtop mLCI system, and concluding with a clinically verified dual- modality instrument.

Optical performance was thoroughly explained and calibration measurements were presented to show linearity and accuracy.

Tissue phantoms were used throughout the development of the instrument for simulations.

Finally, preliminary in vivo human data showed feasibility of the instrument in measuring microbicide gel thickness distributions.

Chapter

4 will now describe the application of this instrument in a clinical study.

It begins with a description of an in vivo study, in which the dual- modality optical imaging instrument was used to measure coating thickness distributions of a placebo microbicide gel distributed along the vaginal lumen.

The study included nine participants that completed fifteen study sessions.

Further details of the clinical study design are provided in Section 4.2.

Data analysis and important summary statistics of the study, such as fraction of tissue surface with coating are included in the following section, Section 4.3.

The results of the study are presented with a comparison of measured distributions between the two modalities.

The results of this study show that 68 the dual- modality optical imaging instrument is capable of providing details of microbicide gel coating in the human vagina.

This data can then be used in biophysical models for objective computations of gel performance.

4.2 Study design Preliminary proof- of- concept in vivo results of the dual- modality device were provided in Section 3.2.3.

Here, it is further demonstrated that the instrument is capable of measuring gel distributions in a human in vivo study, and the quality and utility of the data is evaluated.

The study compares mLCI vaginal gel thickness measurements with simultaneous, co- registered measurements obtained by the fluorimetric technique (Fluorimetric technique is described in Section 3.2.2).

The study consisted of nine participants that completed fifteen study sessions, with some participants completing multiple study sessions.

The study followed a protocol approved by the Institutional Review Board (IRB) at the Duke University Medical Center.



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