Workshop Abstracts
Spectral flow Workshop
Simon Monard (Walter and Eliza Hall Institute of Medical Research)
Alexis Gonzales (The University of Melbourne)
Intro to spectral flow cytometry:
- Why go full spectrum?
- How is it different from conventional flow cytometry
- Challenges and opportunities
- What do we mean “unmixing errors”
- Staining
Optimising reference controls:
- Including autofluorescent signatures
- How bright is too bright
- Group specific negatives
- Beads or cells.
Trouble shooting:
- Why does my data look weird?
- Is it staining or unmixing?
- Post acquisition compensation; A necessary evil?
- Examples of odd looking data and how it was “fixed”.
We will start the workshop with an overview of full spectrum cytometry, how it differs from conventional cytometry and its similarities. The experimental workflow will be demonstrated and the term “reference controls” will be introduced. The term “unmixing errors” is often heard. It is implied that due to a shortcoming of the instrument or the software the data doesn’t meet the users expectations. Unmixing errors are usually due to 1. Inappropriate reference controls, 2. autofluorescence, 3 Poorly designed experiments or 4. staining issues eg. failing to block unwanted staining.
We will explore reference control issues and when should one include one or more autofluorescence signatures as “colours” in the experiment and demonstrate how this works in practice.
Finally, we want to show some examples of experiments where the data looks terrible and how and if such data can be “fixed”. When is it permissible to alter compensation post unmixing and when is this falsifying data by distorting it to meet our expectations. We would like to discuss this last part with the audience.
Sorting Fact from Fiction Tips For Evaluating Your Next Cell Sorter
Eva Orlowski-Oliver (Peter MacCallum Cancer Centre)
Purchasing a new fluorescence-activated cell sorter (FACS) is one of the most significant investments a Shared Resource Laboratory (SRL) can make. With multiple platforms now offering advances in sensitivity, automation, spectral technologies, biosafety features, and operational efficiencies, selecting the most appropriate instrument requires far more than reviewing specifications and vendor demonstrations.
This workshop will provide SRL professionals with a practical framework for evaluating and comparing cell sorting platforms before procurement. Participants will share their own experiences and explore key technical, operational, and strategic considerations that should inform purchasing decisions, including instrument performance, sort purity and recovery, viability, population resolution, biosafety requirements, workflow integration, service support, throughput demands, and other additional costs.
Through real-world examples and interactive discussion, attendees will explore experiments and validation studies that can be used to objectively assess competing instruments. The workshop will also address practical considerations such as facility infrastructure, user requirements, future scalability, and stakeholder engagement.
By the end of the session, participants will have an understanding of how to move beyond marketing claims and generate the evidence needed to make informed, defensible purchasing decisions. This workshop is intended for SRL management, flow cytometry specialists, and institutional decision-makers involved in the acquisition, evaluation, or replacement of cell sorting platforms.
Analysis of Spatial Transcriptomics and CITE-seq Data
A/Prof Alen Faiz (University of Technology Sydney)
The emergence of spatial transcriptomics and Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE-seq) has enabled unprecedented characterization of cellular phenotypes and tissue organization. While CITE-seq provides high-resolution transcriptomic and protein-level characterization of individual cells, spatial transcriptomics preserves the tissue context necessary to understand cellular interactions and microenvironmental structure.
This workshop will focus on analytical strategies for spatial transcriptomic and CITE-seq datasets. Participants will be introduced to workflows for multimodal data preprocessing, quality control, dimensionality reduction, , and cell-type annotation using both transcriptomic and protein-derived information. The workshop will demonstrate methods for constructing robust cellular reference atlases from CITE-seq data and projecting these cell states onto spatial transcriptomic datasets.
Advanced analyses will include identification of spatially variable genes, characterization of cellular neighbourhoods and spatial co-localization analyses. Participants will also explore approaches for identifying disease-associated cellular niches, quantifying tissue architecture, and linking molecular programs to specific anatomical regions. Particular emphasis will be placed on integrating protein and transcriptomic information to improve cell-state resolution and enhance spatial mapping accuracy.
Hands-on examples will demonstrate the use of contemporary computational frameworks in R , including workflows implemented in Seurat and complementary spatial analysis programs. The workshop will highlight best practices for data integration, interpretation of results, and common analytical pitfalls encountered when working with large-scale single-cell and spatial datasets.
By the end of the workshop, participants will have gained practical experience in CITE-seq and spatial transcriptomic data, enabling them to identify cellular populations.
The Grey Zone: Interactive Case-Based Challenges in Ambiguous Lineage Leukaemias
Dr Mary Sartor, BSc, MSc, PhD
Dr Julie Curtin
Accurate classification of acute leukaemias with ambiguous lineage remains one of the greatest diagnostic challenges in flow cytometry. Overlapping immunophenotypic features, aberrant antigen expression, and evolving classification criteria often require the integration of flow cytometry with morphology, cytogenetics, molecular findings, and clinical context. These cases can challenge even the most experienced laboratory professionals and frequently benefit from multidisciplinary discussion.
This interactive workshop will present a series of six to eight cases that illustrate the diagnostic dilemmas encountered in ambiguous lineage leukaemias, including mixed phenotype acute leukaemia (MPAL) and other diagnostically challenging entities. Participants will be actively involved in the decision-making process through live audience polling using QR code technology. For each case, attendees will review the available data and vote on the most likely diagnosis or next diagnostic step via multiple-choice questions, with the aggregated results displayed in real time.
Following each poll, Dr Julie Curtin will lead a discussion exploring the rationale behind the correct interpretation, highlighting key immunophenotypic features, potential pitfalls, and practical approaches to challenging cases. A dedicated question-and-answer session will follow, allowing further exploration of complex issues raised during the workshop.
This session is designed for laboratory scientists, haematopathologists, clinicians, and trainees with an interest in diagnostic flow cytometry.
Cytometry for Clinical Immunophenotyping
Melissa Nardi
Peter MacCallum Cancer Centre
A/Prof Helen McGuire
University of Sydney
Dr. Kevin Lo
CellCarta
This workshop will cover practical approached to implementing spectral cytometry in a clinical setting. Spanning discussions on workflows, validation, and certification considerations, we will be showcasing data problems … as well as how to fix them!
Viable CD34 testing principles
The Royal College of Pathologists of Australasia – Quality Assurance Programs
Workshop description
This workshop focuses on viable CD34 testing principles. The presentations will focus on ISHAGE guidelines, the importance of pre and post analytical variables, common pitfalls and quality matters. The session will include time for case scenarios and panel discussions.
Session 1: Technical component
- Speaker 1
- ISHAGE-based approach with viability
- Single-platform vs dual-platform considerations
- Gating strategies incorporating viability dyes
- Speaker 2
- Common pitfalls:
- Debris and non-specific binding
- Low event counts
- Compensation and fluorescence spillover
- Comparison of viability dyes:
- 7-AAD
- Propidium iodide
- Amine-reactive dyes
- Pros/cons in:
- Fresh samples
- Cryopreserved / thawed samples
- Common pitfalls:
- Speaker 3
- Finish with case scenarios/discussion
**Break**
Session 2: Quality + pre and post analytical variables
- Speaker 4
- Cryopreservation and thawing impact
- Result reporting:
- Units
- Significant figures
- Reporting
- Speaker 5
- ACS/national guideline on CD34 enumeration
- EQA
Navigating the world of scientific publishing — a tutorial for early-career researchers in the field of cytometry
Bartek Rajwa (Purdue University)
Helen McGuire (University of Sydney)
Introduction
Publishing is the currency of a research career, yet most PhD students and postdocs learn its rules through trial, error, and inevitable rejection. This tutorial will introduce the full publication process: choosing where to submit, preparing a competitive manuscript, surviving peer review, and understanding what happens after acceptance. It will help early-career researchers make informed, strategic decisions about sharing their work. It will also map a publication landscape that has changed profoundly over the last decade: the rise of open access and preprints, article processing charges and funder mandates, the spread of predatory journals, the impact of bibliometric evaluation, and the new ground rules for generative AI. Cytometry and single-cell science will serve as a running example (including reporting standards and the OMIP article type), but the skills are transferable across the life sciences.
Learning objectives
By the end of this tutorial, attendees will be able to:
1. Understand the scholarly publishing ecosystem and the life cycle of a manuscript, from submission and editorial triage through peer review, decision, revision, and publication, and distinguish the main peer-review models.
2. Compare open-access and subscription models (gold, green, hybrid, diamond) and understand how APCs, licenses, and funder mandates such as Plan S affect where and how they can publish.
3. Recognize predatory and questionable journals using concrete red flags and trusted tools (Think. Check. Submit., DOAJ, COPE, indexing status).
4. Interpret bibliometric indicators (Journal Impact Factor, CiteScore, h-index, altmetrics), understand their limitations, and apply responsible-assessment principles (DORA, CoARA) when judging journals and researchers.
5. Build a publication strategy that matches a study to the right venue and the right audience, choosing among article types (including field-specific formats such as OMIPs).
6. Write an effective cover letter and assemble a complete, compliant submission package.
7. Communicate professionally with reviewers, Associate Editors, and the Editor-in-Chief: preparing point-by-point responses, rebuttals, and appeals, and handling rejection constructively.
8. Apply core research-integrity standards, including authorship/contributorship, conflicts of interest, data and image integrity, reproducibility (FAIR data, RRIDs), and current expectations for disclosing AI use.
Content
This tutorial moves from the big picture to the practical aspects of publishing, with a focus on the needs of researchers who use cytometry. We begin with the publishing landscape and how a paper travels from submission to publication, then examine open access and the economics of publishing — APCs, who pays, equity, and funder mandates. Next we cover how to spot predatory and hijacked journals and how to read bibliometric scores critically: why the Journal Impact Factor is a poor proxy for the quality of an individual paper or researcher, and what other metrics can and cannot tell you. The second half is hands-on: developing a publication strategy and targeting the right audience (with a particular focus on cytometry), writing a cover letter, assembling the submission, and communicating effectively with editors and reviewers through revision and rebuttal. We close with research integrity and reproducibility in the AI era, and the opportunities that Cytometry Part A offers to serve the community as a reviewer or contributing editor.
Intended audience
This session is designed for PhD students, postdocs, and other early-career researchers across the life sciences who are beginning to publish their own work related to cytometry and single-cell science. It will also benefit group leaders mentoring trainees through publication and anyone considering a role as a reviewer, associate editor, or guest editor.
Presenters
The tutorial is led by Bartek Rajwa and Helen McGuire, the Editor-in-Chief and an Associate Editor, respectively, of Cytometry Part A, the official journal of the International Society for Advancement of Cytometry (ISAC).
Bartek Rajwa is a Professor of Computational Biology at Purdue University. His research applies statistical modeling, machine learning, and signal-processing approaches to single-cell and high-dimensional biological measurements — from the mathematics of spectral unmixing and cytometry data analysis to multi-omics integration and reproducible computational workflows for biomedical and microbiological applications. Bartek co-authored over 160 scientific papers.
Helen McGuire is an Associate Professor in the School of Medical Sciences (Faculty of Medicine and Health) at the University of Sydney, where she leads research identifying immune signatures in chronic human diseases using high-dimensional and mass cytometry. Helen is an ISAC Marylou Ingram Scholar (the society’s program for developing the next generation of leaders in cytometry) and has served on the ISAC Council. She is a co-author of more than 120 research publications.
Tips, Tricks and Best Practices Before the Cytometer: Sample Preparation Best Practices and Workflow Efficiency Hacks
Suat Dervish (Garvan Institute of Medical Research)
Erica Vine (Westmead Institute for Medical Research)
- Sample preparation
- Tissue digestion insights
- Wetlab best practice
- Optimising cytometry staining approaches
- Obvious, hidden and dangerous efficiency hacks in the cytometry facility