optimizing tissue microarrays and area/polygon tiles for efficient large-scale imaging

<meta charset="utf8"></head><body>使用 TSAI 加速组织微阵列成像的 MIBI 运行设置

Multiplexed ion beam imaging (MIBI) is a technique to image 40+ proteins simultaneously on histologic tissue sections at up to 250 nm resolution1,2,3. After a histologic tissue section is stained using antibodies tagged with isotopically pure elemental metals, the MIBI instrument performs secondary ion mass spectrometry to simultaneously quantify all the isotopes - and thus expression of all 40+ antigens - at individual spots on the tissue. Performed across grids of millions of spots, the resulting 40+ plex images of protein expression enable the delineation of cell boundaries and identification of specific cell types while preserving spatial context1,2,3,4. This technique has been used by hundreds of users at roughly 20 sites to study the cellular composition, metabolic profiles, and/or architecture of dozens of tissue types as part of examining the immune response to tumors, tissue inflammation caused by infectious agents, neuropathology of dementia, and immune tolerance in pregnancy5,6,7,8,9,10,11.
A key bottleneck in MIBI instrument operation is setting up fields of view (FOVs) - 200&#160;x 200 &#181;m2 to 800&#160;x 800 &#181;m2 areas of the tissue - for imaging. The MIBI images one FOV at a time, up to 800&#160;x 800 &#181;m2, thus imaging larger areas requires stitching multiple FOVs together. Imaging a tissue microarray (e.g., eight circular tissues in Figure 1A) involves placing multiple FOVs spaced apart. To set up FOVs, the manufacturer interface provides 1) an optical camera image of the slide with a crosshair that roughly corresponds to the specified imaging coordinate (Figure 1A) and 2) a secondary electron detector (SED) image that shows the exact area at the coordinate, reportedly accurate to within 0.1 &#181;m (Figure 1B). First, the user roughly positions a single FOV using the optical image. Because the image resolution is only about 60 &#181;m per pixel, if the placement is off by two pixels (2 pixels&#160;x 60 &#181;m per pixel), a standard 400 &#181;m FOV will be off by 30%. Thus, the user must use the SED image to fine-tune the position - a tedious sequence of a dozen steps involving multiple popup windows, typing coordinates into text boxes, slowly nudging the SED with directional control buttons, and often even writing down coordinates on paper (Supplementary Figure 1). This process must be repeated for each spot of a 100+ core tissue microarray (TMA). Some third-party tools can help with the initial rough positioning12. However, they still require some programming knowledge, and final positioning is still done through the dozen-step process. It is also highly troublesome to position grids of adjacent FOVs, which will be later stitched together into a tiled panoramic image.
Thus, the tile/SED/array Interface (TSAI) was developed with the goal of enabling users to rapidly position large numbers of FOVs using an intuitive, interactive graphical interface. TSAI consists of two main components: 1) A web-based graphical user interface (web UI) for rapidly placing TMA points and tissue tiles, and 2) Integrations into the MIBI user control interface for generating a tiled SED image and adjusting FOV positions. If only using the optical image, many FOVs can be roughly positioned and then quickly adjusted using the FOV navigation/adjustment tools (Figure 2, TSAI, left branch). However, if the SED tiling is performed, FOVs can be accurately positioned on the tiled SED image without needing further adjustments in SED mode (Figure 2, TSAI, right branch). Of general interest to hundreds of current MIBI users, these tools make tiling and TMA positioning very simple even for novices and reduce complex MIBI run setups from several hours to a few dozen minutes.

<meta charset="utf8"></head><body>作者聚焦：介绍用于组织成像中快速视场定位的 Tile/SED/Array 接口

使用 Tile/SED/Array 接口在多路复用离子束成像显微镜上快速构建组织微阵列和平铺面积成像 

优化组织微阵列和面积/多边形平铺以实现高效的大规模成像

optimizing-tissue-microarrays-areapolygon-tiles-for-efficient-large

Research

JoVE Journal

Biology

Optimizing Tissue Microarrays and Area/Polygon Tiles for Efficient Large-Scale Imaging

15 次观看. 要为组织微阵列或 TMA 点网格设置视野或 FOV，请打开平铺的 SED 图像。在 tiles 列的相关磁贴中，调整 columns 和 rows。然后选中或取消选中地图中的框。在相关拼贴中，单击 TMA 以打开 TMA 选项 菜单。设置 TMA 污点的行数和列数。如有必要，请添加命名前缀并编辑起始行和列编号。在幻灯片图像上，单击 TMA 的四个角。然后单击并拖动带圆圈的...

视频: 优化组织微阵列和面积/多边形平铺以实现高效的大规模成像

Rapid Setup of Tissue Microarray and Tiled Area Imaging on the Multiplexed Ion Beam Imaging Microscope Using the Tile/SED/Array Interface

Multiplexed ion beam imaging (MIBI) is a next-generation mass spectrometry-based microscopy technique&#160;that generates 40+ plex images of protein expression in histologic tissues, enabling detailed dissection of cellular phenotypes and histoarchitectural organization. A key bottleneck in&#160;operation occurs when users select the physical locations on the tissue for imaging. As the scale and complexity of MIBI experiments have increased, the manufacturer-provided interface and third-party tools have become increasingly unwieldy for imaging large tissue microarrays and tiled tissue areas. Thus, a web-based, interactive, what-you-see-is-what-you-get (WYSIWYG) graphical interface layer - the tile/SED/array Interface (TSAI) - was developed for users to set imaging locations using familiar and intuitive mouse gestures such as drag-and-drop, click-and-drag, and polygon drawing. Written according to web standards already built into modern web browsers, it requires no installation of external programs, extensions, or compilers. Of interest to the hundreds of current MIBI users, this interface dramatically simplifies and accelerates the setup of large, complex MIBI runs.

Multiplexed ion beam imaging (MIBI) is often used to image tissue microarrays and tiled, contiguous tissue areas, but current software for setting up these experiments is cumbersome. The tile/SED/array interface is an intuitive, interactive graphical tool developed to dramatically simplify and accelerate MIBI run setup.

Multiplexed ion beam imaging (MIBI) is a next-generation mass spectrometry-based microscopy technique&#160;that generates 40+ plex images of protein ...