Details

Name
Ubiquitin Cortex Analysis
IQbot ID
IQBOT-0010
Version
1.0.0
Research Area(s)
Cancer Research, Drug Toxicity & Metabolism, Genetics, Neurology, Common Tools
Processing Time Expectation
Less than 2 Minutes per Image
Acquisition Hardware
Wide-Field, Optical Microscope in Bright-Field (Transmission) Mode Using a Color Mosaic CCD or Monochrome CCD with RGB Filter
Techniques, Dyes, Stains, & Proteins
Ubiquitin, 3,3'-Diaminobenzidine (DAB), Horesradish Peroxidase (HRP), Hematoxylin Counterstain, Immunohistochemistry, Chromogen, Brown Oxidation Reaction
Sample Context
In Tissue
Name
Ubiquitin Cortex Analysis
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IQbot ID
IQBOT-0010
Version
1.0.0
Research Area(s)
Cancer Resarchh, Drug Toxicity & Metabolism, Genetics, Neurology, Common Tools
Processing Time Expectation
Less than 2 Minutes per Image
Acquisition Hardware
Wide-Field, Optical Microscope in Bright-Field (Transmission) Mode Using a Color Mosaic CCD or Monochrome CCD with RGB Filter
Techniques, Dyes, Stains, & Proteins
Ubiquitin, 3,3'-Diaminobenzidine (DAB), Horesradish Peroxidase (HRP), Hematoxylin Counterstain, Immunohistochemistry, Chromogen, Brown Oxidation Reaction
Sample Context
In Tissue

Output Parameters

Total Tissue Area (pixels)
The total area of tissue within the image.
Positive Nuclei Count
The number of DAB positive (i.e., brown) regions within the segmented tissue, independent of size.
Positive Nuclei Count
The total area of DAB-positive pixels across the segmented tissue.
Negative Nuclei Area (pixels)
The total area of DAB-positive pixels across the segmented tissue.
Negative Nuclei Area (pixels)
The total sum of hematoxylin-positive pixels (blue, DAB-negative) within the segmented tissue.

 

This section will identify any output parameters that are only output by the IQbot in its advanced configuration.

 

Total Tissue Area (pixels)
The total area of tissue within the image.
Positive Nuclei Count
The number of DAB positive (i.e., brown) regions within the segmented tissue, independent of size.
Positive Nuclei Count
The total area of DAB-positive pixels across the segmented tissue.
Negative Nuclei Area (pixels)
The total area of DAB-positive pixels across the segmented tissue.
Negative Nuclei Area (pixels)
The total sum of hematoxylin-positive pixels (blue, DAB-negative) within the segmented tissue.

Image Acquisition


This IQbot was developed to analyze cortex tissue images acquired using a wide-field, optical microscope in bright-field (transmission) mode using a color mosaic CCD or monochrome CCD with RGB filter. For accurate analysis of tissue area, sections must be counterstained with hematoxylin.

 
 

The Ubiquitin Cortex Analysis IQbot has been built to accommodate the following imaging types only.

 24-bit RGB
 .tif
  .tiff
  .bmp
 .jpg
.jpeg
 
 

As we reference for IQbots in general here, the Ubiquitin Cortex Analysis is only ultimately effective if you upload high quality images for processing. Please ensure that any images you upload follow the below guidelines specific to this IQbot.

High-Resolution:  The quality of the output data you receive is largely dependent on the input image resolution. Please provide high-resolution images to ensure that the IQbot can most effectively differentiate between features.
In-Focus:  Please produce the most in-focus images possible for analysis. Images with high background contrast will result in more accurate and useful data and overlays.
Properly Saturated:  If your submitted images are significantly undersaturated or oversaturated, it is possible that the IQbot will be unable to detect structures, or may group structures together. Please ensure that the images you upload are saturated appropriately.
Clear of Artifacts:  Please take care to avoid creating bubbles, debris, or otherwise inducing artifacts into your image. Such additional features may corrupt the resulting data.
Clear of Accessary Information:  Please do not submit images retaining any annotations, legends, timestamps, or other markings. Every part of the images that you upload will be analyzed and included in the output data.
 
 
 

If you are uncertain whether or not your images meet the quality standards for which this IQbot was validated, or have additional questions relating to image quality, please view our FAQs or contact the ImageQuantify.com support team.

 

     (216) 678-9258           help@imagequantify.com

 

Validation Information


This IQbot has been successfully validated for use, and a copy of its validation certificate accompanies the results of each order it analyzes. In accordance with general ImageQuantify.com validation procedures, this IQbot has been found to provide quantitative information equivalent to that of a manual observer, but with no variability and increased breadth of information.

 
 

The first step of the validation process was to establish the ground truth against which the IQbot’s efficacy would be compared. For the purpose of the Ubiquitin Cortex Analysis IQbot, ground truth was defined on a manual observer basis. Trained observers were guided through a standardized, semi-automated process to produce output data from a specially selected validation image set. This image set represented the entire range of expected output values (e.g., different configurations, labeled targets, and Ubiquitins densities). Images were presented for analysis in a randomized order and deidentified to protect against observer biases. The observer mean for each parameter measured was accepted as ground truth.

By analyzing the inter- and intraobserver variability present in the manual observation process, acceptable criteria were established so as to compare the IQbot’s output to that of a trained replacement observer. Ultimately, the IQbot’s performance proved to offer significant upgrades over manual or semi-automated analysis techniques, and validation was considered successful. Further considerations ensured that the IQbot rejected and flagged concerning image-specific outputs and performed within operation time constraints.

 

The Ubiquitin Cortex Analysis IQbot identifies ubiquitin (DAB reporter) positive cells and nuclei in the cortex of brain histological sections counterstained with hematoxylin. Ubiquitin is regulatory molecule that is responsible for tagging cell proteins for proteasomal destruction. Thus ubiquitin presence in cortical neurons could be used to characterize brain health and study progression of various neurological diseases. This IQbot processes each stained histological cross-section image and extracts the total tissue coverage (Total Tissue Area), number of DAB or ubiquitin positive cells (Positive Nuclei Count), total area of DAB or Ubiquitin present in the tissue (Positive Nuclei Area), number of DAB negative nuclei (Negative Nuclei Count) and total area of DAB negative (hematoxylin positive) nuclei within the tissue (Negative Nuclei Area). Additionally, the IQbot also generates two pseudo-colored overlay images which contain (1) a red outline delineating the tissue area/boundary, blue regions indicating ubiquitin negative nuclei, and green areas depicting ubiquitin positive cells (*_overlay1.tif) and (2) a red outline delineating the tissue area/boundary and green areas depicting ubiquitin positive cells (*_overlay2.tif).

DEMO IMAGE
IQBOT APPLIED IMAGE

 

SAMPLE DATA
Image Total Tissue Area
(pixels)
Positive Nuclei Count Positive Nuclei Area
(pixels)
Negative Nuclei Count Negative Nuclei Area
(pixels)
Sample 1062716 531 11155 6280 93825