Articles

Protocol optimization is essential for clear, reproducible results   Fluorescence microscopy uses fluorophore-labeled antibodies and other fluorescent reagents to visualize targets of interest in cells and tissues. Experiments can be as simple as detecting one...

  Understanding the biological density of proteins or antigens expressed by each individual cell is an imperative component of all cell-based analytical methods. The comparative change in protein expression, also called antigen density, is indicative of the...

Reducing autofluorescence is critical in fluorescence-based research   Techniques such as fluorescence microscopy, flow cytometry, and western blotting often rely on the use of fluorophore-labeled antibodies. The main reason for this is that fluorescence-based...

Fluorescent cellular analytical technologies allow us to “see” beyond what was historically possible with histological stains or morphological scatter profiles. In the early days, microscopy employed excitation sources like arc lamps, isolating specific wavelengths of...

Choosing the right fluorophores is critical for reliable results   Fluorescent detection offers significant advantages, including multiplexing capability, superior sensitivity, and a broader dynamic linear range compared to  other detection methods. However, the...

Understanding the advantages and disadvantages of different immunodetection strategies is important to achieve accurate results.   Microscopy-based techniques such as immunocytochemistry (ICC) and immunohistochemistry (IHC) often use fluorophore or enzyme-labeled...

When designing a flow cytometry experiment, it is important to account for the relative brightness of each fluorescent label on your specific instrument.  Ideally, brighter fluorophores should be assigned to weakly expressed markers, while dimmer fluorophores should...

Advanced microscopy techniques enable deeper imaging   Advanced microscopy platforms are becoming more widespread for the depth of information they provide. Among these newer modalities, confocal microscopy has risen in popularity for imaging thick...

Whether preformulated or made in-house, buffers are integral to flow cytometry success   Flow cytometry allows researchers to characterize individual cells using fluorophore-labeled antibodies for detecting targets of interest. Sample preparation, antibody...

Non-conventional optics and advanced algorithms are extending the capabilities of classic microscopy.   For centuries, researchers have tried to improve the resolution and sensitivity of optical microscopy. Over time, advances in fluorescence imaging modalities...

Flow cytometry controls must address multiple sources of variation   Flow cytometry requires more controls than other immunoassay techniques because it accounts for a greater number of potential sources of variation. In addition to experimental controls that...

The commercial availability of antibody-linked fluorescent dyes has expanded rapidly over the past decade. Where researchers were once limited to a handful of common dyes (FITC, PE, APC, PerCP, etc.), they can now choose from an extensive list of fluorescent dye lines...

Fluorescence recovery after photobleaching is a technique for characterizing the mobility of cellular molecules   Fluorescence recovery after photobleaching (FRAP), also known as fluorescence redistribution after photobleaching, is a microscopy-based technique...

Antibody conjugate panels must be designed with care to ensure accurate results Immunophenotyping is a flow cytometry-based technique which uses fluorophore-labeled antibodies for identifying different cell types within a heterogeneous population. While some cell...

Microscopy is comprised of many innovative technologies and remains at the forefront of scientific research.   Since the invention of the microscope in the late 16th century, there has been a continual push to produce instruments with higher resolution, faster...

Fixation and permeabilization are key protocol steps for several core immunoassay techniques Immunoassay techniques such as flow cytometry, immunocytochemistry (ICC), and immunohistochemistry (IHC) often require that samples be fixed and/or permeabilized prior to...

The use of spectral flow cytometry is increasing due to the many benefits it affords. Spectral flow cytometry is a relatively new technology that was developed to push the boundaries of traditional flow cytometry. First described in 2004 by researchers at Purdue...

Near-infrared fluorophores offer several advantages for imaging applications. A major advantage of fluorescent detection is that it enables multiple antibodies to be combined in the same experiment. Using techniques such as immunocytochemistry (ICC) and...

Fluorescence microscopy is seeing increased utility for imaging organisms While fluorescence microscopy has long been employed for imaging plated mammalian cells, its use for visualizing organisms is flourishing. In recent years, winning images in Nikon’s annual Small...

High viabilities, well-preserved antigens, and minimal clumping underpin flow cytometry success Unlike bulk population analysis techniques such as ELISA and Western blot, flow cytometry provides information about individual cells. Although microscopy does this as...

What are OMIPs?  Optimized multicolor immunofluorescence panels, commonly known as OMIPs, are highly validated sets of antibodies and reagents that are published, peer-reviewed, and compiled to serve as a resource to the scientific community. OMIP publications often...

New approaches to Western blotting techniques overcome performance limitations For over 40 years, researchers have used Western blot for protein identification. Not only does Western Blot provide confirmation that a target of interest is present in a sample, this...

Expansion Microscopy physically expands biological specimens enabling higher resolution imaging   First described in 2015, expansion microscopy is an imaging technique that improves on the resolution of conventional light microscopy by physically expanding the...

Identifying distinct stem cell types and monitoring their differentiation is essential for developing novel therapies   Stem cell research originated in the 1950s when it was discovered that mice which had received a lethal dose of radiation could recover...

Excluding dead cells from flow cytometry analysis with a cell viability fluorescent dye is critical for producing accurate results   The presence of dead cells in samples that will be analyzed by flow cytometry can affect antibody staining and lead to inaccurate...

Compared to traditional monolayers, 3D cell cultures more closely resemble physiological conditions 2-dimensional (2D) cell cultures consisting of monolayers attached to a flat surface have been a mainstay of scientific research for decades. However, cells grown in...

Spectral flow cytometry offers several advantages over conventional flow cytometry but may be more affected by spreading error Spectral flow cytometry was first described in 2004 by the Robinson group at Purdue University Cytometry Laboratories. Since then, several...

Confocal microscopy offers several advantages over traditional widefield microscopy and has evolved to meet many different research needs Originally developed by Marvin Minsky in the 1950s to address the problem of light scatter, confocal microscopy has become an...

Failure to correctly identify cell-cell complexes can lead to data being misinterpreted and compromise the purity of sorted cell populations Multiparametric flow cytometry allows researchers to detect different cell types within a heterogeneous population. When...

Choosing the right antibody for the intended application and sample type is critical to producing reliable results Antibodies are essential tools for applications ranging from flow cytometry, immunocytochemistry (ICC), and immunohistochemistry (IHC) to Western...

Software packages for viewing and analyzing FCS data files offer different features and benefits Data analysis represents a major source of variation in flow cytometry. Not only is the analysis process highly prone to user subjectivity, but disparities may arise due...

Fluorescent proteins have enabled countless scientific discoveries, but they have their limitations For decades, researchers have been fascinated by the ability of certain living organisms to emit light. However, it was not until 1962 that this phenomenon was first...

Repurposing the CRISPR-Cas system for imaging genomic targets in living cells Established methods for cell imaging use fluorophore-labeled antibodies or oligonucleotides to bind proteins or nucleic acids in fixed cells or rely on the engineered expression of...

Compensation is one of the most important but least understood aspects of multicolor flow cytometry Multicolor flow cytometry allows researchers to characterize complex cellular populations by staining for several cell type specific markers at the same time. However,...

Advantages of TSA include increased sensitivity, simplified panel design, and faster time to results Tyramide signal amplification (TSA), also known as catalyzed reporter deposition (CARD) or tyramine amplification technique (TAT), is a method used for detecting...

Flow cytometry continues to evolve with advances in instrumentation, reagents, and software The first fluorescence-based flow cytometer was developed in 1968 and was commercialized the following year. Since then, the field of flow cytometry has matured rapidly, and...

Studying calcium flux leads to insights into normal physiology and disease Calcium ions (Ca2+) are involved in numerous intracellular signaling cascades and in a broad range of physiological processes. These include neurotransmission, muscle contraction, and hormone...

Marker selection and panel design underpin the accuracy of cellular identification Being able to monitor the presence and relative abundance of different cell types in sample material is essential to understand normal biology and investigate the pathogenesis of...

Aberrant cell proliferation and cell death underlie a multitude of disease states Normal tissue homeostasis depends on a critical balance between cell proliferation and cell death. The cell cycle regulates the former, while the latter occurs via controlled...

 Confocal microscopy is a powerful imaging technique used to study biological specimens. It offers several advantages over conventional widefield microscopy, including the capacity to control depth-of-field and collect serial sections from thick samples. Fluorophores...