Multiplexing in fluorescence light microscopy

Semester: Spring
|
Year offered: 2026

Course description

This 7-hours long (over 2 days) nanocourse introduces the principles and applications of multiplexing in light microscopy, a powerful approach to visualize multiple molecular targets within the same sample. Through lectures (day 1) and hands-on activities (day 2), participants will learn the rationale for multiplexing, explore iterative and non-iterative technologies, and understand the advantages and limitations of each method. The course will cover and emphasize practical considerations such as experimental design, antibody validation, labeling strategies, panel design, and hardware requirements. Additionally, the course will cover key aspects of image acquisition, optimization, and analysis, including 2D/3D registration, unmixing, and visualization resources. In the lab sessions, participants will gain hands-on experience with SpectraPlex, FLIM-based multiplexing, linear unmixing, and data visualization platforms, using both sample data and real microscopy experiments. By the end of the course, participants will be equipped with practical skills and knowledge to design, execute, and analyze multiplexed imaging experiments in their own research.

Session dates, times, and location

Day 1 (TMEC 128 Learning Studio): Tuesday, February 17th, 1:30-4:30PM 

Day 2 (MicRoN Facility, the exact room will be communicated directly by the instructor on Day 1 of class):

  • Group A: Wednesday, February 18th, 1:30–5:30 PM
  • Group B: Friday, February 20th, 1:30–5:30 PM

 

Prerequisites

This is an entry-level course. No previous knowledge of the topic is required, as the course addresses the fundamentals of multiplexing. 

 

Course objectives 

By the end of this course, participants will be able to:

  • Define multiplexing in light microscopy and explain its applications.

  • Compare iterative and non-iterative multiplexing technologies, with their pros and cons.

  • Design multiplexed experiments, including antibody validation, labeling, and panel selection.

  • Identify hardware and software requirements for multiplexed imaging.

  • Visualize and analyze multiplexed datasets using available computational resources.

 

Milestone credit 

In order to receive a Milestone, students must:

  • Attended both sessions for the entire time.

  • Complete a course evaluation final survey.

More information about Milestone Credit can be found here.

 

Course Team

Course director/instructor

Paula Montero Llopis, PhD, MicRoN Director,paula_monterollopis@hms.harvard.edu

Praju Vikas Anekal, PhD, Adrienne Wells, PhD, Han Su, PhD, Clarence Yapp, PhD

Curriculum Fellow

Lorenzo Gesuita, PhD, Lecturer on Genetics, Harvard Genetics Department, Lorenzo_Gesuita@hms.harvard.edu

 

Enrollment limit 

A maximum of 32 students will be allowed. Priority will be given to Harvard graduate students taking the course for credit. Harvard Postdocs can register too, and they will be granted access to the course as space allows.

 

Registration link & deadline

Please register using this link by Friday, February 6th, 2026:
https://hms.az1.qualtrics.com/jfe/form/SV_6Ja1anqw1gLt410 

See also: