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AutoTag and AutoSnap: Standardized, semi-automatic capture of regions of interest from whole slide images.

Marien KM, Andries L, De Schepper S, Kockx MM, De Meyer GR - MethodsX (2015)

Bottom Line: Although SURS is the most reliable sampling method, it implies a high workload.However, SURS can be semi-automated and in this way contribute to the development of a validated quantification method for microvessel counting in the clinical setting.Here, we report a method to use semi-automated SURS for microvessel counting: •Whole slide imaging with Pannoramic SCAN (3DHISTECH)•Computer-assisted sampling in Pannoramic Viewer (3DHISTECH) extended by two self-written AutoHotkey applications (AutoTag and AutoSnap)•The use of digital grids in Photoshop(®) and Bridge(®) (Adobe Systems) This rapid procedure allows traceability essential for high throughput protein analysis of immunohistochemically stained tissue.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Physiopharmacology, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium ; HistoGeneX NV, ZNA Middelheim Campus, Lindendreef 1, B-2020 Antwerp, Belgium.

ABSTRACT
Tumor angiogenesis is measured by counting microvessels in tissue sections at high power magnification as a potential prognostic or predictive biomarker. Until now, regions of interest (ROIs) were selected by manual operations within a tumor by using a systematic uniform random sampling (SURS) approach. Although SURS is the most reliable sampling method, it implies a high workload. However, SURS can be semi-automated and in this way contribute to the development of a validated quantification method for microvessel counting in the clinical setting. Here, we report a method to use semi-automated SURS for microvessel counting: •Whole slide imaging with Pannoramic SCAN (3DHISTECH)•Computer-assisted sampling in Pannoramic Viewer (3DHISTECH) extended by two self-written AutoHotkey applications (AutoTag and AutoSnap)•The use of digital grids in Photoshop(®) and Bridge(®) (Adobe Systems) This rapid procedure allows traceability essential for high throughput protein analysis of immunohistochemically stained tissue.

No MeSH data available.


Related in: MedlinePlus

Snapshots of regions of interest (ROIs) combined with a counting grid of a stained slide can be produced semi-automatically and with full traceability. After the stained slide has been scanned in the Pannoramic SCAN (3DHISTECH), it needs to be checked manually for possible image quality issues (out-of-focus regions, incomplete images) in Pannoramic Viewer (3DHISTECH). In the next step, point annotations are semi-automatically created in the whole slide image with AutoTag. Standardized snapshots of the regions of interest (ROIs) can now be automatically created with AutoSnap in Pannoramic Viewer. Finally, a digital grid in Photoshop and Bridge (Adobe Systems) is combined with the snapshots of the ROIs.
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fig0005: Snapshots of regions of interest (ROIs) combined with a counting grid of a stained slide can be produced semi-automatically and with full traceability. After the stained slide has been scanned in the Pannoramic SCAN (3DHISTECH), it needs to be checked manually for possible image quality issues (out-of-focus regions, incomplete images) in Pannoramic Viewer (3DHISTECH). In the next step, point annotations are semi-automatically created in the whole slide image with AutoTag. Standardized snapshots of the regions of interest (ROIs) can now be automatically created with AutoSnap in Pannoramic Viewer. Finally, a digital grid in Photoshop and Bridge (Adobe Systems) is combined with the snapshots of the ROIs.

Mentions: Tumor tissue was stained for CD31 (NCL-CD31-1A10, Leica Biosystems, Diegem, Belgium) on a Ventana Benchmark® XT platform in an accredited laboratory (HistoGeneX NV, Antwerp, Belgium). The stained slides were scanned with the slide scanner Pannoramic SCAN (3DHISTECH) to obtain a whole slide image (Fig. 1). A 20× Plan Apo objective (0.80 NA3) and Hitachi (HV-F22CL) 3CCD progressive scan color camera with a resulting image resolution of 0.23 μm/pixel were used. JPEG image encoding with quality factor 80 and an interpolated focus distance of 15 with stitching in the scan options were chosen. For every slide a specific scan profile was configured and holes in the scan area were filled to allow for correct detection of tissue and in-focus images of the tissue. Scanned images were examined to check for image quality and to confirm that the whole tissue section was captured (Fig. 2A).


AutoTag and AutoSnap: Standardized, semi-automatic capture of regions of interest from whole slide images.

Marien KM, Andries L, De Schepper S, Kockx MM, De Meyer GR - MethodsX (2015)

Snapshots of regions of interest (ROIs) combined with a counting grid of a stained slide can be produced semi-automatically and with full traceability. After the stained slide has been scanned in the Pannoramic SCAN (3DHISTECH), it needs to be checked manually for possible image quality issues (out-of-focus regions, incomplete images) in Pannoramic Viewer (3DHISTECH). In the next step, point annotations are semi-automatically created in the whole slide image with AutoTag. Standardized snapshots of the regions of interest (ROIs) can now be automatically created with AutoSnap in Pannoramic Viewer. Finally, a digital grid in Photoshop and Bridge (Adobe Systems) is combined with the snapshots of the ROIs.
© Copyright Policy - CC BY
Related In: Results  -  Collection

License
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getmorefigures.php?uid=PMC4487922&req=5

fig0005: Snapshots of regions of interest (ROIs) combined with a counting grid of a stained slide can be produced semi-automatically and with full traceability. After the stained slide has been scanned in the Pannoramic SCAN (3DHISTECH), it needs to be checked manually for possible image quality issues (out-of-focus regions, incomplete images) in Pannoramic Viewer (3DHISTECH). In the next step, point annotations are semi-automatically created in the whole slide image with AutoTag. Standardized snapshots of the regions of interest (ROIs) can now be automatically created with AutoSnap in Pannoramic Viewer. Finally, a digital grid in Photoshop and Bridge (Adobe Systems) is combined with the snapshots of the ROIs.
Mentions: Tumor tissue was stained for CD31 (NCL-CD31-1A10, Leica Biosystems, Diegem, Belgium) on a Ventana Benchmark® XT platform in an accredited laboratory (HistoGeneX NV, Antwerp, Belgium). The stained slides were scanned with the slide scanner Pannoramic SCAN (3DHISTECH) to obtain a whole slide image (Fig. 1). A 20× Plan Apo objective (0.80 NA3) and Hitachi (HV-F22CL) 3CCD progressive scan color camera with a resulting image resolution of 0.23 μm/pixel were used. JPEG image encoding with quality factor 80 and an interpolated focus distance of 15 with stitching in the scan options were chosen. For every slide a specific scan profile was configured and holes in the scan area were filled to allow for correct detection of tissue and in-focus images of the tissue. Scanned images were examined to check for image quality and to confirm that the whole tissue section was captured (Fig. 2A).

Bottom Line: Although SURS is the most reliable sampling method, it implies a high workload.However, SURS can be semi-automated and in this way contribute to the development of a validated quantification method for microvessel counting in the clinical setting.Here, we report a method to use semi-automated SURS for microvessel counting: •Whole slide imaging with Pannoramic SCAN (3DHISTECH)•Computer-assisted sampling in Pannoramic Viewer (3DHISTECH) extended by two self-written AutoHotkey applications (AutoTag and AutoSnap)•The use of digital grids in Photoshop(®) and Bridge(®) (Adobe Systems) This rapid procedure allows traceability essential for high throughput protein analysis of immunohistochemically stained tissue.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Physiopharmacology, University of Antwerp, Universiteitsplein 1, B-2610 Antwerp, Belgium ; HistoGeneX NV, ZNA Middelheim Campus, Lindendreef 1, B-2020 Antwerp, Belgium.

ABSTRACT
Tumor angiogenesis is measured by counting microvessels in tissue sections at high power magnification as a potential prognostic or predictive biomarker. Until now, regions of interest (ROIs) were selected by manual operations within a tumor by using a systematic uniform random sampling (SURS) approach. Although SURS is the most reliable sampling method, it implies a high workload. However, SURS can be semi-automated and in this way contribute to the development of a validated quantification method for microvessel counting in the clinical setting. Here, we report a method to use semi-automated SURS for microvessel counting: •Whole slide imaging with Pannoramic SCAN (3DHISTECH)•Computer-assisted sampling in Pannoramic Viewer (3DHISTECH) extended by two self-written AutoHotkey applications (AutoTag and AutoSnap)•The use of digital grids in Photoshop(®) and Bridge(®) (Adobe Systems) This rapid procedure allows traceability essential for high throughput protein analysis of immunohistochemically stained tissue.

No MeSH data available.


Related in: MedlinePlus