\section{Results} %1200 words\cite{EileenJBO,SierraCDAblation,AnthonyFO2015}
Video-mosaicking is of particular interest for rapid assessment of skin lesions that can be arbitrarily large and/or located at curved, irregular anatomical sites. We illustrate the capabilities of our approach by presenting results for three distinct clinical cases,  each of which presents a particular  set of imaging requirements. The flexibility and robustness of our approach is demonstrated for producing useful video-mosaics in all three settings.The first case is a lentigo maligna melanoma (LMM), where the need for accurate planning of surgical margins (Sec. \ref{LMM_Surg}) at non-flat corrugated and irregular anatomical sites is met by video-mosaicking over arbitrarily large predefined areas of tissue. In the second case, we show how video-mosaicing can help in adaptively detecting small and focal sites of suspicion within widely spread areas for diagnosing and monitoring of extramammary  Paget disease (Sec. \ref{Paget}). In the third  case, we illustrate how video-mosaicking can help in rapid intraoperative detection of residual basal cell carcinoma in surgical wounds (Sec. \ref{BCCMargin}). All the videos acquired for this study were obtained after Institutional Review Board approval. 

%\del{In \ref{LMM_Surg}, we show how arbitrarily large predefined areas of tissue can be imaged and displayed. In \ref{Paget}, we show how users can image over large areas of tissue in real-time adaptive fashion. Finally in \ref{BCCMargin}, we show, how clinicians can perform in vivo imaging directly in a surgical wound to rapidly examine arbitrarily shaped residual tumor margins in an adaptive fashion.}\com{repetitive again if placed here, to my eye; maybe we can use this sentence later --- stay tuned !} 

\subsection{Case 1: Lentigo Maligna Melanoma (LMM)}\label{LMM_Surg}
In Figure \ref{Case:LMM}, we illustrate the use of our approach for pre-surgical planning of LMM excisions. LMMs frequently occur on the face, head. and neck. The lesions usually appear to be small on the surface (millimeters in extent), but the sub-clinical sub-surface spread is often substantially larger (centimeters), and is also often obscured within surrounding degraded photo-aged skin. Moreover,  the sub-surface spread of melanoma cells is typically highly dendritic, like the roots of a tree, and therefore the thin tendrils that define the outermost margins are difficult to determine using current approaches, namely visual (clinical, dermoscopic) inspection and Wood’s lamp (ultraviolet-excited fluorescence) examination.

\begin{figure}[ht]
\centering
\includegraphics[width = 0.9\textwidth]{./Fig_LMM.eps}
\caption{\textbf{Lentigo Maligna Melanoma (LMM):} Figure illustrates three potential methods to treat an LMM lesion in the clinical setting and how RCM video-mosaicking can improve LMM management. In all three approaches, following an initial diagnostic biopsy (red asterisk),  the clinical margin (yellow dotted circle in (A,B,C)) is determined through macro imaging (dermoscopy and Wood's lamp examination). (A) In the standard surgical procedure, completely excising cancer in least number of surgery is the primary concern and tissue conservation secondary. After diagnosis, in a single visit, the lesion is excised with a 1 cm wide surgical margin around the clinical margin. (B)  At specialized cancer care clinics, in order to spare healthy tissue, multi-staged excision may be performed. In the case illustrated here, first an excision with a conservative margin (5mm, red circle 1) was conducted. Histological examination of the excised tissue, indicated residual tumor at 11 o'clock spreading in a radial direction. Thus, after a few days, a second excision was planned and executed in that region only (red circle 2). In (C) we show how RCM video-mosaicking can streamline and improve the procedure. The video-mosaic of the clinical margin was created from in-vivo RCM videos (shown overlaid on the clinical image) by the algorithm described here. Expert examination of the video-mosaic revealed tumor extension at the 11 o'clock location and extending radially (yellow inset), therefore, we extended the video-mosaic in that direction (the radial mosaic in the direction of the red arrow, white inset), all in one visit. Histopathological examination along the same line as the extended mosaic (red arrows in panels (D) and (E)) confirmed the findings in the RCM video-mosaic, showing the extent and margin of tumor cell spread towards the periphery of the lesion.}
\label{Case:LMM}
\end{figure}

Following an initial diagnostic biopsy (at the location of the red asterisk at the center of panels (A), (B), and (C)), the clinical margin (yellow dotted circle) of the lesion is determined with dermoscopy and Wood’s lamp examination. In the standard surgical procedure, removal of the disease is the primary concern and tissue conservation the secondary one. Therefore, as shown in Figure \ref{Case:LMM}-(A), a wide “safety” margin (red dotted circle), up to 1cm, beyond the clinical margin is estimated. The estimated margin is interrogated by blindly sampling the margin via ”mapping" biopsies (blue asterisks) \cite{Dengel2008}. Several days later, if all biopsy reports result in negative findings at the margin, the lesion is excised along the surgical margins. In case any of the biopsies turn out to be positive, the surgical margin is extended further along that direction. After the surgery, if residual tumor is found at the edges of the excised tissue, then further surgery is performed on a third visit.
 
In specialized cancer care centers, in order to conserve more of the healthy tissue, alternative multi-stage excision guided by histology is sometimes performed (Figure \ref{Case:LMM}-(B). After the initial diagnostic biopsy, the central part of the lesion corresponding to the clinical margin (yellow dotted circle) plus an initial narrower “safety” margin (usually 5 mm) is excised (red dotted circle). After histopathological analysis of the excised tissue, if it turns out that the excision margin in any quadrant contains residual tumor, the patient undergoes further excision in those positive  quadrant(s) (red dotted curve) and the excised tissue is processed again by pathology. This process iterates until tumor-free  margins are achieved and can take several days to several weeks, depending on the size and irregularity of the tumor.

In contrast, our video-mosaicking  method offers a streamlined diagnostic and therapeutic procedure, as illustrated  in Figure \ref{Case:LMM}-(C). Sub-surface structures are visualized non-invasively  along the clinical  margin (yellow  dashed circle),  as shown in the circular video-mosaic superimposed in gray-scale at the clinical margin.  Assessment of the video-mosaic revealed tumor in one quadrant (yellow inset) and consequently a localized radial scan was performed next (illustrated in detail in \ref{Case:LMM}-(D)). Expanded views of the yellow and white insets show two frames along this radial scan. Analysis of the radial mosaic correctly determined the extent of the tumor growth (without the need for mapping biopsies) in the associated quadrant. of the tissue to guide further surgery. Immediately after imaging, the lesion was excised with an appropriate ”safety” margin (red dotted lines). Note that the capability of RCM for sub-surface imaging with cellular resolution allows non-invasive detection of the extent of disease, which  otherwise is not possible without invasive biopsy.

Histopathology from this particular case confirmed the margin determined  in the video-mosaic. Note that he white arrows in panel D show two locations at which our tailored video-mosaicking algorithm automatically compensates for "jumps" (discontinuities) in the video, allowing correct mosaicking of the three distinct arcs. The individual  sub-mosaics are put together manually in an image editor. This is a simple procedure as the first and last frame of consecutive sub-mosaics are spatially located next to (or very close to each other), and their relative location to each other can typically be estimated using the RCM video.

The resulting video-mosaicking method allows sufficiently  precise determination of the appropriate surgical margin  \cite{OriolLMM}. The excision is tissue-conserving, similar to that of the staged excision approach used in specialized clinics, but all accomplished in a single visit without  the need for repeated visits and repeated biopsies.


\subsection{Case 2:Extramammary Paget disease (EMPD)} \label{Paget}
In Figure \ref{Case:Pagets}, we illustrate the use of video-mosaicking to improve the sampling accuracy and guide diagnosis of extramammary Paget disease (EMPD). EMPD is a rare skin cancer that appears on the genitalia,  often covering large areas, and that is frequently mistaken for an inflammatory or infectious condition. In addition, due to a patchy and focal distribution of tumor cells, sampling the lesion for diagnosis and management requires numerous biopsies, most of which typically turn out to be benign.

In order to cover such large areas with curved surface topography, clinicians move the confocal microscope over the region of interest in a free-hand  fashion. The approach is to acquire RCM videos in the suspicious area, starting from the center of the lesion and navigating towards the outer borders in a spiral path, aiming to cover the entire area in a rapid fashion without missing any regions. This approach allows identification of areas with higher tumor cell density, which can then be selectively biopsied to confirm diagnosis.\cite{OriolPaget}

Being able to image in a user-determined irregular trajectory is important in EMPD as tumor cells are sparsely distributed in a patchy fashion. To illustrate, in Figure \ref{Case:Pagets}, we indicate a region (yellow asterisk on panels A and B) that is well within the clinical lesion but determined in the video-mosaic to be free of tumor cells. Furthermore, we also illustrate a positive area (white dotted circle in A and B), where video-mosaic suggests signs of EMPD in a clinically suspected area. The findings from video-mosaicking are also confirmed by histological examination (Figure \ref{Case:Pagets}-C) of the biopsied sample from the region. 

\begin{figure}[ht]
\centering
\includegraphics[width = 0.9\textwidth]{./Pagets.eps}
\caption{\textbf{Extramammary Paget Disease (EMPD):} Figure illustrates the use of video-mosaicking for non-invasive examination of tumor spread in a suspected case of EMPD. Panel A (lower left) shows a clinical image exhibiting the suspicious lesion, whose red color and appearance is suggestive of EMPD. In panel B (top), a video-mosaic is obtained by imaging with a handheld confocal microscope in a spiral fashion intended to largely and efficiently cover the green box in panel A. The region inside the white circle on panels A and B was determined to be suspicious in the video-mosaic, and thus a histological section (yellow line in panel B) was excised (shown in panel C). Histological examination of the slice confirmed the findings from the video-mosaic, as illustrated by the high density of tumor cells visible at the yellow arrow in both panels B and C. Unfortunately, during such imaging along a spiral path, the operator could not keep the imaging depth constant and the resulting artifact, as seen in upper left side of the mosaic, occurred at the overlap area. Although the algorithm stitching artifacts through grapcuts based method, such  artifacts cannot be completely avoided due to the nature of RCM imaging and challenges in image capture.}
\label{Case:Pagets}
\end{figure}

\subsection{Case 3: Basal Cell Carcinomas} \label{BCCMargin}
Figure \ref{Case:BCC} illustrates  the use of video-mosaicking for the intraoperative assessment of residual basal cell carcinoma (BCC) margins during surgery. BCC is often treated with a tissue conserving procedure called Mohs micrographic surgery, in which the surgeon is guided by the preparation and reading of frozen pathology of thin excisions, taken in successive stages. This is a relatively slow procedure since each stage takes 20-45 minutes to be processed for frozen sections and 2 or more stages are often required  for clearance.

\begin{figure}[ht]
\centering
\includegraphics[width = \textwidth]{./BCC.eps}
\caption{\textbf{Basal cell carcinoma (BCC):} Video-mosaic showing residual tumor (inset, bright lobular nests) in a quadrant of a Mohs surgical wound. Video-mosaicking enables the localization of the residual tumor by providing architectural information that is not readily available when viewing the original videos.}
\label{Case:BCC}
\end{figure}

Video-mosaicking performed directly in the Mohs surgical wound allows rapid detection of residual tumor.\cite{EileenJBO} This enhances the ease of correlating microscopic cellular detail with macroscopic location in the wound, which would be much more difficult with standard RCM imaging or by simply  observing the original  videos. Since the video-mosaic has the same resolution as in the original images, the surgeon can zoom in on suspicious areas and examine them in detail, with higher magnification, without loss of any information  (e.g., inset in Figure \ref{Case:BCC}).