Immunological Images and the ImmPort Database and Analysis Portal Anna Maria Masci Department of Immunology Duke University Buffalo, 24 June 2014

Multiscale System Immunology project PI Thomas Kepler Responsible for data submission to ImmPort Grace Kepler Patrick Dunn

WHAT DO WE DO WITH DATA Data to run a simulation Biology Computational Data to upload to ImmPort

HISTOLOGY FACS ELISAELISPOT SEQUENCING BIOLOGY

CONTEXT HISTOLOGY PROVIDES INFORMATION IN A CONTEXT

Kindly provided by Luigi Racioppi

Histology Image ABOUT Anatomical entity DEFINED by molecule expression DETECTED by stain VISIBLE by light emission CAPTURED by microscopy STORE as image

PMID: LZ: GL7, CD83, CD86 DZ: GL7, CD83, CD86 Markers versus Location

Victora, GD. Science 9 August 2013: vol. 341 no

Victora, GD. Science 9 August 2013: vol. 341 no

PMID: Structure of germinal centers sign of Sjogren syndrome

input transformation output Organism Organ Tissue Cell Reagent Assay Assay Objective Treatment Instrument Data format Process data Statistical Analysis General approach to experimental methods

INPUT Organism Organ Tissue Cell Molecule UBERON, FMA, MA CL PRO, CheBi Automatic? Mouse Lymph node follicle germinal center B cell GL7 Ability to extract data about germinal center B cell by querying for data about mouse lymph node that use reagent GL7 Ontology can facilitate the automatic query

Reagents and Materials Solutions, antibodies OBI REO Antibody repository Properties of the reagents And material PH Concentration Specificity Glass Plastic Slide, plate

Anti mouse IgM is_a Rabbit Immunoglobulin has_label Texas Red recognizes mouse Immunoglobulin isotype M has_quality monoclonal

TRANSFORMATION Assay Assay Target Treatment Instrument OBI BARD BAO Confocal microscopy Intra-vital microscopy Lymph node Surgery

Instrument Type of instrument Settings Type of detection Limit of detection Zeiss LSM 510 confocal 488, 543, and 633 nm excitation lines Plan-Apochromat 20× (NA 0.75) Nikon NIS-Elements Imaging Software Objective Fluorescence emission

OUTPUT Data format Processed data Statistical analysis Histogram, image, excel Germinal center structure, protein expression value Intensity: min, max, median, sum, std. dev., and center on a per channel basis Statistic analysis output Comma Separate Value, Excel

IMMPORT BASIC STUDY DESIGN PROTOCOLS SUBJECTS BIOSAMPLE REAGENTS EXPERIMENTS TREATMENT

REAGENTS Indicated in blue are fields we believe should be added to the ImmPort template Reagent role: is about the role the reagent is playing in the assay. E.g. dilution role if the reagent is use to dilute another reagent Reagent concentration: is about the working concentration of a reagent. Differences in reagent concentrations affect the result Reagent specificity: is about what a reagent is recognizes. E.g. anti mouse IgM is specific for a mouse IgM

PROTOCOL Instrument: defines the type of instrument, different instruments have different feature and those are necessary type of information for comparison of data capture, using different type of instruments Microscopy settings: is about the parameters that are set during image capturing by a microscope. Laser different microscope are equipped with different laser that are activated at different wavelength. Activation of different wavelength result in different light emission and this affect the intensity of the image. Magnification: picture with different magnifications are difficult to compare. Type of detection: is about the type of channel use to read the light emission Limits of detection are important because when an instrument is set close to a limit of detection errors are more easily to happen.

5 Histology experiment sample Processed data: a final information value obtained. E.g. protein expression value Data format: In which format is the data? E.g. Histogram, excel. Statistical analysis: which type of statistical analysis has been applied by the instrument or the researcher to get the processed data Experiment target: which is the target of the experiment. E.g Germinal center B cell location Experiment output: is the final type of information you get from the experiment. E.g. spatial coordinates Section depth: is about the thickness of the slide and impact the resolution Section series: it is important to know if an image is or is not part of a series, if it is is important to know which number is in a series.

PMID: transcription profiling identification objective cellular feature identification objective Investigation Germinal Center Formation Investigation Germinal Center Formation

Victora Lab HP_ Inability to walk SNOMEDCT/ Walking