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Welcome to our beginner's guide to multiplex technologies for antibody-based applications. Multiplexing is a powerful technique that allows researchers to simultaneously measure multiple parameters within a single experiment. This efficient and data-rich approach is revolutionizing various fields of biological and medical research. On this page, you'll find essential information about the basics of antibody-based multiplex technologies, including flow cytometry, western blot, imaging, and immunoassays. Whether you're new to these methods or looking to expand your knowledge, this guide will provide you with the foundational understanding you need to get started.
The benefit of multiplexing
Multiplexing techniques offer numerous advantages over traditional singleplex experiments. The ability to simultaneously analyze multiple targets within a single sample improves efficiency and productivity while helping reduce costs and enhancing data quality. When investigating complex biological pathways, it offers the opportunity to gain a deeper understanding of how a biological system responds to external stimuli. The simultaneous detection of multiple protein targets in one experiment helps reduce experimental variability and is an excellent choice when the sample volume is limited or when analyzing large sample sets. Selecting the right products from the start and careful experimental planning and design helps reduce time spent on optimization and troubleshooting and helps ensure the success of your experiment.
Advantages of multiplex versus singleplex
- Increased efficiency—conduct multiple measurements in a single experiment, helping save time and resources
- Cost-effectiveness—reduce the overall cost per data point by minimizing the number of separate experiments required
- Enhanced data richness—obtain comprehensive datasets that help provide deeper insights into complex biological processes
- Reduced sample volume—minimize the amount of sample needed for analysis, which is particularly beneficial when working with precious or limited materials
Choosing the right multiplex technology: A beginner’s guide
Navigating the world of multiplex technologies can be daunting, especially for those who are new to the field. To help you get started, we've created a comprehensive selection table that compares the key aspects of four major multiplex technologies for protein and cell analysis: flow cytometry, western blot, imaging, and immunoassays. This table highlights essential parameters such as the principle of each technology, key applications, sample types, sensitivity, and more. By providing a side-by-side comparison, we aim to give you a clear understanding of the strengths and limitations of each method, enabling you to choose the most suitable technology for your research needs.
Parameter |
Flow cytometry |
Imaging |
Immunoassay |
Western Blot |
Principle |
Measures physical and chemical characteristics of cells/particles |
Visualizes and quantifies cellular/ subcellular structures; detects morphological changes |
Detects and quantifies specific biomolecules |
Detects specific proteins in a sample via antibody binding |
Key applications |
Cell sorting, protein expression, cell health |
Cellular localization, morphology studies, cell health analysis |
Biomarker detection, disease diagnostics |
Protein detection, expression analysis |
Sample type |
Cells, tissues, microorganisms, and other biomolecules |
Fixed and live cells, tissues, multiwell plates, 3D models (e.g., spheroids, organoids) |
Serum, plasma, cell lysates |
Proteins, cell and tissue lysates |
# of targets |
1–40 |
1–8 |
1–80 |
1–5 |
Sensitivity |
High |
High |
High |
Medium to high |
Quantitative |
Yes |
Yes |
Yes |
Yes |
Required sample volume |
100 µL of sample if the cell concentration is sufficiently high (1–10 million cells/mL) |
EVOS Systems: Varies based on slide or dish size. CX7 Systems: 50–100 µL per well for 96-well plates. 6.3 µL per well for 384-well plates |
25–50 µL for 96-well 6.3 µL for 384-well |
From a few µL to up to 100 µL |
Data analysis |
Attune Software, FlowJo, FCS Express |
Celleste Image Analysis Software, |
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Panel design tools |
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Build your own multiplexing panels
To help simplify your research, we offer a suite of intuitive online tools designed to help you create custom multiplexing panels with ease. Whether you're working with flow cytometry, western blot, imaging, or immunoassays, our tools provide step-by-step guidance to help ensure you select the right markers and reagents for your experiments. Enhance your research efficiency and accuracy by leveraging our powerful online tools and take your experiments to the next level.
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