Learn about a wide range of applications and techniques related to using primary and secondary antibodies in your cell biology research. Topics include immunocytochemistry, immunohistochemistry, immunofluorescence, immunophenotyping using flow cytometry, and western blot analysis.
Sensitive immunodetection in cells and tissues requires high-affinity antibodies that exhibit minimal background staining. Invitrogen Superclonal secondary antibodies represent a breakthrough in recombinant antibody technology, providing sensitive binding to their targets and very low levels of nonspecific staining.
Both differentiated and undifferentiated stem cells are identified and characterized using primary antibodies to key targets, which are detected by fluorescence-based methods such as immunofluorescence imaging and flow cytometry. This article features the characterization of new antibodies for stem cell research.
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| Type | Title | Categories |
|---|---|---|
| Application note (2011) | Six-color mouse lymphocyte immunophenotyping using the Attune Acoustic Focusing Cytometer with red laser option | Alexa Fluor, antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, multicolor flow cytometry, particles |
| Application note (2011) | Immunophenotyping peripheral blood cells: A no-lyse, no-wash, no cell loss method for immunophenotyping nucleated peripheral blood cells using the Attune Acoustic Focusing Cytometer | antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, flow cytometry sample preparation, fluorescent dyes, immunophenotyping, particles |
| Application note (2011) | Mouse immunophenotyping: A no-lyse, no-wash, and no cell loss method using the Attune Acoustic Focusing Cytometer | antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, flow cytometry sample preparation, fluorescent dyes, immunophenotyping, particles |
| Application note (2011) | Detection of phosphoproteins using the Attune Acoustic Focusing Cytometer | Alexa Fluor, antibodies, Attune/Attune NxT, cell signaling, flow cytometer/flow cytometry, fluorescent dyes, protein detection |
| Application note (2011) | Detection of human mesenchymal stem cells using the Attune Acoustic Focusing Cytometer | Attune/Attune NxT, cell cycle, flow cytometer/flow cytometry, immunophenotyping, stem cell research |
| Application note (2012) | Detecting human circulating endothelial cells using the Attune Acoustic Focusing Cytometer | Alexa Fluor, antibodies, Attune/Attune NxT, endothelial cells, flow cytometer/flow cytometry, fluorescent dyes, rare-event detection |
| Application note (2013) | Optimizing the high content analysis of cell morphology using multi-dimensional imaging on the Thermo Scientific ArrayScan XTI HCA Reader: Stem cell colonies | Alexa Fluor, antibodies, ArrayScan, confocal microscopy, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, immunofluorescence (IF), microplate reader, particles, stem cell research |
| Application note (2013) | Multiplexed mitosis and apoptosis analysis | antibodies, apoptosis, ArrayScan, cell cycle, cell proliferation, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, microplate reader, particles |
| Application note (2015) | Ten-parameter immunophenotyping of human lysed whole blood with the Attune NxT Flow Cytometer allows identification of multiple T cell subsets and myeloid cells | Attune/Attune NxT, flow cytometer/flow cytometry, immunophenotyping, multicolor flow cytometry |
| Application note (2015) | Multiparameter analysis of murine regulatory T cells and dendritic cells with the Attune NxT Flow Cytometer | Attune/Attune NxT, flow cytometer/flow cytometry, immunophenotyping, multicolor flow cytometry |
| Application note (2015) | Detection of murine regulatory T cells on the Attune NxT Flow Cytometer | Attune/Attune NxT, flow cytometer/flow cytometry, immunophenotyping, multicolor flow cytometry |
| Application note (2015) | Attune NxT Flow Cytometer for 6-Color immunophenotyping analysis of stained human whole blood using a no-lyse, no-wash protocol, with no compensation | Attune/Attune NxT, flow cytometer/flow cytometry, immunophenotyping, multicolor flow cytometry |
| Application note (2015) | Quantitation of proliferating cells with the EVOS FL Auto Imaging System | antibodies, antibody labeling, ArrayScan, cell proliferation, EVOS FL Auto Microscope, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, onstage incubator, phagocytosis |
| Application note (2015) | No-wash, no-lyse detection of phagocytic cells via a pHrodo BioParticles functional assay in human whole blood on the Attune NxT Flow Cytometer | antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, flow cytometry sample preparation, fluorescent dyes, immune system, immunophenotyping, particles, pH detection |
| Application note (2015) | 13-parameter immunophenotyping of human lysed whole blood with the the Attune NxT Flow Cytometer allows for the identification of B cells, NK cells, multiple T cell subsets and myeloid cells | Alexa Fluor, antibodies, Attune/Attune NxT, cancer, flow cytometer/flow cytometry, fluorescent dyes, immune system, immunophenotyping, multicolor flow cytometry, nanocrystals, Qdot |
| Application note (2016) | Blood cell counting using the Countess II FL Automated Cell Counter | automated cell counter, brightfield microscopy, cell counting, cell counting sample preparation, Countess, EVOS, fluorescent dyes, immunophenotyping, ReadyProbes |
| Application note (2017) | Detection of platelets in whole blood using the Attune NxT Flow Cytometer | antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, fluorescence, immunophenotyping, platelets |
| Application note (2017) | Flow cytometry analysis of transcription factor expression during differentiation of hPSC-derived cardiomyocytes | antibodies, Attune/Attune NxT, cardiomyocytes, cell differentiation, flow cytometer/flow cytometry, fluorescence, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), immunophenotyping, stem cell research |
| BioProbes articles (Issues 50– present day) | BioProbes Journal of Cell Biology Application | cell analysis, flow cytometry, imaging microscopy, immunoassays, antibodies, protein detection and quantification |
| Molecular Probes Handbook | TSA and other peroxidase-based signal amplification techniques—Section 6.2 | Alexa Fluor, antibodies, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), reagents, secondary detection, signal amplification |
| Molecular Probes Handbook | Qdot nanocrystals—Section 6.6 | avidins and streptavidins, cell tracking, fluorescence, particles, Qdot, secondary antibodies |
| Molecular Probes Handbook | Zenon Technology: Versatile reagents for immunolabeling—Section 7.3 | antibody labeling, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, secondary detection, Zenon |
| Molecular Probes Handbook | Secondary immunoreagents—Section 7.2 | antibodies, avidins and streptavidins, secondary antibodies |
| Pathway poster (2012) | TNF signaling pathway | antibodies, cell signaling, immunoassays |
| Pathway poster (2012) | Toll-like receptor signaling pathway | antibodies, cell signaling, immunoassays |
| Pathway poster (2012) | The human kinome | antibodies, cell signaling, immunoassays |
| Pathway poster (2012) | Integrin signaling pathway | antibodies, cell signaling, immunoassays |
| Pathway poster (2012) | Interferon signaling pathway | antibodies, cell signaling, immunoassays |
| Pathway poster (2012) | MAPK signaling pathway | antibodies, cell signaling, immunoassays |
| Pathway poster (2012) | Akt/mTOR signaling pathway | antibodies, cell signaling, immunoassays |
| Pathway poster (2012) | Jak/Stat signaling pathway | antibodies, cell signaling, immunoassays |
| Protocol | Immunofluorescent staining of intracellular antigens on cultured cells | fluorescence microscopy/fluorescence imaging, general ab staining, imaging, immunofluorescence (IF) |
| Protocol | Immunohistochemical staining of formalin-fixed paraffin-embedded tissues | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC frozen tissue—Indirect method (purified) | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC frozen tissue—Indirect method (biotin) | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC frozen tissue—Direct method | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC FFPE Tissue Trypsin Digestion—Direct Method | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC FFPE tissue trypsin digestion antigen retrieval—Indirect method | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC FFPE tissue low pH antigen retrieval—Indirect method | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC FFPE tissue low pH antigen retrieval—Direct method | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC FFPE tissue high pH antigen retrieval—Indirect method | general ab staining, imaging, immunohistochemistry (IHC) |
| Protocol | IHC FFPE tissue high pH antigen retrieval—Direct method | general ab staining, imaging, immunohistochemistry (IHC) |
| Scientific poster (2007) | Evaluation of click chemistry-based alternative to BrdU antibody labeling in tissue and cultured cells using fluorescence microscopy and flow cytometry | antibodies, ArrayScan, cell cycle, cell proliferation, Click-iT, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, fluorescent dyes, microplate reader, nucleic acid labeling |
| Scientific poster (2007) | Click chemistry-based detection of S-phase adherent cells using automated microscopy and image analysis | Alexa Fluor, antibodies, ArrayScan, cell cycle, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, microplate reader |
| Scientific poster (2007) | Click catalyzed nucleic acid labeling as a novel replacement for BrdU antibody-based cell proliferation assay | Alexa Fluor, antibodies, antibody labeling, cell cycle, cell proliferation, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, nucleic acid labeling |
| Scientific poster (2008) | Characterization of DNA content, cyclin B1 and phosphorylated histone H3 with direct S-phase using EDU incorporation in multiparameter testing of cell lines with cell cycle blocking agents | Alexa Fluor, antibodies, apoptosis, ArrayScan, cell cycle, cell proliferation, Click-iT, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis |
| Scientific poster (2008) | Zenon labeling technology facilitates detection of intra-cellular phosphoprotein determinants | Alexa Fluor, antibodies, cell signaling, flow cytometer/flow cytometry, fluorescent dyes, protein labeling, rare cell detection, Zenon |
| Scientific poster (2008) | Novel Click-IT TUNEL Assay for detection of cell death | Alexa Fluor, antibodies, apoptosis, Click-iT, cytotoxicity, fluorescence microscopy/fluorescence imaging, fluorescent dyes, gel electrophoresis, high content analysis, immunocytochemistry (ICC), viability |
| Scientific poster (2008) | Detection of S-phase cell cycle progression using 5-ethynyl-2'-deoxyuridine incorporation with click chemistry | Alexa Fluor, antibodies, ArrayScan, cell cycle, Click-iT, flow cytometer/flow cytometry, fluorescent dyes, high content analysis, microplate reader, nucleic acid labeling, nucleic acid quantitation |
| Scientific poster (2008) | Detection of DNA synthesis by automated microscopy and image analysis: Comparison of BRDU method and a new click chemistry-based EDU method | Alexa Fluor, antibodies, ArrayScan, cell cycle, cell proliferation, Click-iT, flow cytometer/flow cytometry, fluorescent dyes, high content analysis, microplate reader, nucleic acid labeling |
| Scientific poster (2009) | A comparative study of 25 different CD4 conjugates | Alexa Fluor, antibodies, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, multicolor flow cytometry, nanocrystals, Qdot |
| Scientific poster (2009) | Proliferative and phenotypic characterization of human mesenchymal stem cells by flow cytometry | Alexa Fluor, cell proliferation, Click-iT, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, nucleic acid quantitation, stem cell research, violet laser-excited reagents |
| Scientific poster (2009) | Dead cell stains in flow cytometry: A comprehensive analysis | apoptosis, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, viability |
| Scientific poster (2009) | Qdot nanocrystal conjugates in multispectral flow cytometry | Alexa Fluor, antibodies, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, multicolor flow cytometry, nanocrystals, Qdot, violet laser-excited reagents |
| Scientific poster (2009) | Quantitative analysis of genotoxicity and cytotoxicity to DNA damaging agents using high-content imaging | Alexa Fluor, antibodies, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, nucleic acid labeling, nucleic acid quantitation, viability |
| Scientific poster (2010) | Acoustic flow cytometry for multicolor immunophenotyping analysis | antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, fluorescent dyes, immune system, immunophenotyping, multicolor flow cytometry, nanocrystals, Qdot, violet laser-excited reagents |
| Scientific poster (2010) | Acoustic focusing cytometry: Sensitivity and throughput | apoptosis, Attune/Attune NxT, cell cycle, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, phagocytosis |
| Scientific poster (2010) | High content imaging of autophagy | antibodies, ArrayScan, autophagy, BacMam technology, cell signaling, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, live-cell imaging, microplate reader |
| Scientific poster (2010) | Antibody- and fluorescent protein-based approaches to measuring autophagy in mammalian cells by fluorescence microscopy | Alexa Fluor, antibodies, autophagy, BacMam technology, cell proliferation, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, live-cell imaging |
| Scientific poster (2010) | Proliferative and phenotypic characterization of human mesenchymal stem cells by flow cytometry and imaging | Alexa Fluor, cell proliferation, Click-iT, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, fluorescent dyes, immunofluorescence (IF), immunophenotyping, stem cell research, violet laser-excited reagents |
| Scientific poster (2010) | Autophagosomal accumulation perturbs Golgi structure without affecting other organelles: Implications for autophagosome biogenesis | Alexa Fluor, antibodies, autophagy, BacMam technology, cell structure-golgi, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, live-cell imaging |
| Scientific poster (2010) | Use of surface-modified microspheres for setting compensation in multicolor flow cytometry | antibodies, flow cytometer/flow cytometry, flow cytometry compensation, fluorescent dyes, multicolor flow cytometry, particles, violet laser-excited reagents |
| Scientific poster (2010) | ABfinity antibodies: A novel approach to antibody development | Alexa Fluor, antibodies, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, fluorescent dyes, gene expression, immunocytochemistry (ICC), western blotting |
| Scientific poster (2010) | A comparison of three techniques to induce efficient ex vivo T cell expansion | Alexa Fluor, antibodies, cell proliferation, CellTrace, flow cytometer/flow cytometry, fluorescent dyes, immune system, magnetic beads, multicolor flow cytometry, viability |
| Scientific poster (2010) | Identification and characterization of O-GlcNAc modification of galectin-1 in mesenchymal stem cells using click chemistry | Alexa Fluor, BacMam technology, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, gel electrophoresis, immunocytochemistry (ICC), protein enrichment, protein labeling, western blotting |
| Scientific poster (2010) | Rare event detection using acoustic cytometry | Attune/Attune NxT, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, rare-event detection |
| Scientific poster (2011) | Acoustic flow cytometry for multicolor immunophenotyping analysis | antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, multicolor flow cytometry, nanocrystals, Qdot, violet laser-excited reagents |
| Scientific poster (2011) | No-lyse no-wash immunophenotyping using acoustic cytometry | Attune/Attune NxT, flow cytometer/flow cytometry, flow cytometry sample preparation, fluorescent dyes, immunophenotyping, nanocrystals, Qdot |
| Scientific poster (2011) | Acoustic cytometry for rare event detection of PNH cells | Attune/Attune NxT, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, rare-event detection |
| Scientific poster (2011) | Simultaneous analysis of cell death mechanisms and oxidative stress using high content imaging | Alexa Fluor, antibodies, apoptosis, ArrayScan, autophagy, caspase substrates, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, microplate reader, oxidative stress |
| Scientific poster (2011) | Primary human cells from Life Technologies: Offerings, scale and applications in HCS and HTS formats | Alexa Fluor, antibodies, autophagy, BacMam technology, cell proliferation, cell signaling, Click-iT, drug discovery, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis |
| Scientific poster (2011) | Validation of high content imaging and analysis assays for cell health and cytotoxicity | antibodies, apoptosis, ArrayScan, autophagy, caspase substrates, cell health, cell proliferation, CellLight, Click-iT, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, microplate reader, pH detection |
| Scientific poster (2011) | The next step in the analysis of lymphocyte proliferation | Alexa Fluor, antibodies, Attune/Attune NxT, cell proliferation, CellTrace, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, live-cell imaging, violet laser-excited reagents |
| Scientific poster (2011) | Flash & Glow 1,2-dioxetane chemiluminescent substrate formulations for clinical research platforms | antibodies, chemiluminescence, ELISA, enzyme activity detection, magnetic beads, microplate reader, substrate comparison |
| Scientific poster (2011) | The next generation of cell-based imaging assays for apoptosis, autophagy and oxidative stress from Molecular Probes | Alexa Fluor, antibodies, apoptosis, ArrayScan, autophagy, caspase substrates, fluorescence microscopy/fluorescence imaging, fluorescent dyes, high content analysis, microplate reader, oxidative stress |
| Scientific poster (2011) | Site-specific click chemistry-mediated labeling of antibody glycans using metabolic and enzymatic approaches | Alexa Fluor, antibodies, antibody labeling, Click-iT, fluorescent dyes, western blotting |
| Scientific poster (2012) | Studying malaria using the Attune flow cytometer | Attune/Attune NxT, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, malaria, parasite identification |
| Scientific poster (2012) | High sensitivity applications for the Applied Biosystems Attune | Attune/Attune NxT, caspase substrates, flow cytometer/flow cytometry, flow cytometry sample preparation, fluorescent dyes, immunophenotyping, viability |
| Scientific poster (2012) | A novel no-wash, no-lyse method for residual leukocyte enumeration using acoustic focusing flow cytometry and a cell-membrane permeant DNA dye | Attune/Attune NxT, cell counting, flow cytometer/flow cytometry, flow cytometry sample preparation, fluorescent dyes, immunophenotyping, rare-event detection, violet laser-excited reagents |
| Scientific poster (2013) | Pacific Green dye: A new fluorophore for violet laser excitation | Alexa Fluor, antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, fluorescent dyes, immunophenotyping, multicolor flow cytometry, nanocrystals, Qdot, viability, violet laser-excited reagents |
| Scientific poster (2013) | Improved click chemistry demonstrating EdU cell proliferation with GFP-expressing cells and R-PE-based immunophenotyping | Alexa Fluor, Attune/Attune NxT, cell cycle, cell proliferation, Click-iT, flow cytometer/flow cytometry, fluorescent dyes, fluorescent proteins, immunophenotyping |
| Scientific poster (2013) | Cells coming to life: Tools for visualizing immune response | Attune/Attune NxT, endocytosis/phagocytosis, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, immunophenotyping, live-cell imaging, pH detection |
| Scientific poster (2015) | Colorimetric dual labeled EdU/BrdU technology demonstrates contextual information and dynamics of proliferation in tissue | apoptosis, brightfield microscopy, cell proliferation, Click-iT, brightfield microscopy, fluorescence microscopy/fluorescence imaging, immunohistochemistry (IHC) |
| Scientific poster (2015) | A no lyse, no-wash approach to characterizing phagocyte phenotype and function in whole human blood on the Attune NxT Flow Cytometer | Attune/Attune NxT, cell cycle, flow cytometer/flow cytometry, immunophenotyping, no lyse, phagocytosis, whole blood |
| Scientific poster (2016) | Correlating internalization and potency to accelerate antibody discovery and development | brightfield microscopy, colorimetric, endocytosis and phagocytosis, EVOS,fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), immunohistochemistry (IHC) |
| Scientific poster (2016) | Cell mediated cytotoxicity in "untouched" whole blood | Attune/Attune NxT, cell concentration, cell cycle, flow cytometer/flow cytometry, immunophenotyping, no lyse, whole blood |
| Scientific poster (2016) | No-lyse no-wash new strategy for CD34+ for absolute cell counting without beads | antibodies, Attune/Attune NxT, flow cytometer/flow cytometry, flow cytometry compensation, immunophenotyping, multicolor flow cytometry, particles |
| Scientific poster (2016) | Incorporating histological analysis into existing high content imaging | antibodies, ArrayScan, brightfield microscopy, fluorescence microscopy/fluorescence imaging, high content analysis, immunohistochemistry (IHC) |
| Scientific poster (2016) | A microsphere-based universal compensation approach | Attune/Attune NxT, Flow cytometer calibration, flow cytometer/flow cytometry, flow cytometry sample preparation, immunophenotyping, multicolor flow cytometry |
| Scientific poster (2016) | Click colorimetric EdU proliferation and TUNEL: Click chemistry for brightfield microscopy | apoptosis, brightfield microscopy, cell proliferation, Click-iT, brightfield microscopy, fluorescence microscopy/fluorescence imaging, immunohistochemistry (IHC) |
| Tutorial | 1.2 Fixation–Fixed cell imaging: 5 steps for publication-quality images The next step of tissue preparation is fixation. Fixation refers to a chemical means of killing and preserving cells in a particular physiological state, and in many cases, to preserve morphology. Proper fixation equals preservation of target. | fixation, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF) |
| Tutorial | 4.1 Mounting media–Fixed cell imaging: 5 steps for publication-quality images Using the right mounting media can impact your experiment. Be sure to choose the right type of mountant for your set-up. | antifade mounting media, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), mounting media |
| Tutorial | 3.2 Secondary antibody optimization–Fixed cell imaging: 5 steps for publication-quality images Secondary antibody detection protocols also need to be optimized for each primary antibody used. | antibody labeling, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), secondary antibodies, secondary antibody protocol, secondary detection |
| Tutorial | 4.2 Photobleaching and antifades–Fixed cell imaging: 5 steps for publication-quality images What is photobleaching and how can you prevent it from destroying your sample? Options for antifades are discussed. | antifades, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), photobleaching, Prolong Gold, Vectashield |
| Tutorial | 5.5 Ethical considerations–Fixed cell imaging: 5 steps for publication-quality images Ethical imaging means trustworthy data, and thus, publishable data. How to treat your samples and data to preserve data integrity is presented. | ethical considerations, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, image analysis, immunofluorescence (IF), sample manipulation |
| Tutorial | 5.2 Imaging platforms-software–Fixed cell imaging: 5 steps for publication-quality images Taking images on a microscope usually entails having some type of imaging software that aids in taking the image and assists in combining differing colors into one. There are some very important aspects to keep in mind to get a publishable image. | FiFI software, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, ImageJ, imaging software, imaging systems, immunofluorescence (IF) |
| Tutorial | 1.1 Culture conditions–Fixed cell imaging: 5 steps for publication-quality images The first step in obtaining a good image is tissue preparation. For cultured cells, the cells must have good cell health and morphology, as well as good confluency. Healthy cells equal healthy data. | EVOS, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, imaging hardware, imaging systems, immunofluorescence (IF), microscope |
| Tutorial | 1.5 Autofluorescence–Fixed cell imaging: 5 steps for publication-quality images The last step in cell preparation is autofluorescence. Cells and tissue can have a certain degree of autofluorescence that can confuse the specific signal, and lower the signal-to-background. Overcoming autofluorescence means greater sensitivity. | autofluorescence, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), tissue autofluorescence |
| Tutorial | 5.4 Image analysis with Celleste software–Fixed cell imaging: 5 steps for publication-quality images The functionality of the Celleste softwis reviewed and considerations for processing the image in different software programs are described. | Celleste software, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, image analysis, immunofluorescence (IF) |
| Tutorial | 1.4 Blocking–Fixed cell imaging: 5 steps for publication-quality images The next step after permeabilization is blocking, and there are a number of blocking techniques. Protein blocking equals specific antibody binding. Dye charge blocking means less non-specific binding. | dye charge blocking, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), nonspecific binding, protein blocking |
| Tutorial | 2.2 Primary antibody protocol optimization–Fixed cell imaging: 5 steps for publication-quality images Every primary antibody must be optimized separately. There are many protocols available, and it is important to understand a "one size fits all" approach gives inferior results, as every antibody is slightly different. Learn how to approach optimization. | antibody labeling, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), primary antibodies, primary antibody protocol |
| Tutorial | 5.1 Imaging platforms-hardware–Fixed cell imaging: 5 steps for publication-quality images The fifth step of the process is the actual imaging. To capture top-quality images, you need an imaging platform with top-of-the-line imaging capabilities. Here we review considerations for getting the best image. | EVOS, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, imaging hardware, imaging systems, immunofluorescence (IF), microscope |
| Tutorial | 3.5 Dye choice and special concerns–Fixed cell imaging: 5 steps for publication-quality images There are many different dyes spanning the visible, far-red, and infrared wavelengths.Considerations for making the right choices for your experiment are presented. | dye choice, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, Fluorescence SpectraViewer, immunofluorescence (IF), label choice, multiparametric dye selection |
| Tutorial | 1.3 Permeabilization–Fixed cell imaging: 5 steps for publication-quality images The next step is the permeabilization of the cells which is the key to opening intracellular compartments. | cell permeabilization, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF) |
| Tutorial | 3.4 Controls–Fixed cell imaging: 5 steps for publication-quality images Researchers should conduct all necessary controls to rule out the possibility of non-specific binding or non-specific signal. Types of controls are described. Proper controls will boost your confidence in your final results. | fixed-cell imaging, fluorescence microscopy/fluorescence imaging, imaging controls, immunofluorescence (IF), sample controls |
| Tutorial | 5.3 Image capture with EVOS FL Auto 2.0–Fixed cell imaging: 5 steps for publication-quality images Here the advantages of using the EVOS FL Auto 2.0 imaging system to capture your images are discussed. | EVOS FL Auto 2.0, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, image capture, immunofluorescence (IF) |
| Tutorial | 3.3 Amplification techniques–Fixed cell imaging: 5 steps for publication-quality images If the signal is not strong enough using standard secondary detection schemes, you can increase the signal using amplification techniques. This is particularly important for low-expressing antigens, or rare-cell detection in samples. Examples are discussed. | biotin and streptavidin, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), signal amplification, tyramide detection |
| Tutorial | 2.1 Primary antibody choice–Fixed cell imaging: 5 steps for publication-quality images After preparation, the second step to publishable images is to label the sample, usually involving primary antibodies to your specific targets of interest. The antibody source, the use of direct versus labeled antibodies as well as the validation for specific applications is discussed. | antibody labeling, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), primary antibodies |
| Tutorial | 3.1 Secondary antibody choice–Fixed cell imaging: 5 steps for publication-quality images Step three of the five steps in making publishable images is to detect the label. That is, to detect with a secondary antibody, for instance, or an amplification technique, as well as to determine what controls to use. Your options are discussed. | antibody labeling, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), secondary antibodies, secondary antibody, secondary detection |
| Tutorial | Labeling a purified antibody: Educational video on how to label your next antibody for imaging or fluorescence Walk through the tutorial as Molecular Probes scientists demonstrate the protocol—including all the tips and tricks you'll want to know about for your next antibody labeling experiment. The video features Judie showing Curtis, a chemistry graduate student, how to label his monoclonal antibodies while saving time and maximizing yield. | antibodies, antibody labeling, fluorescence microscopy/fluorescence imaging, fluorescent dyes, immunofluorescence (IF) |
| Video | How to site specifically label your antibody using SiteClick technology This video explains how to easily and site specifically label an antibody using an enzymatic and Click chemistry approach. This method can be applied to any intact IgG antibody and requires no antibody engineering or complex methodology. Unlike classic antibody conjugation techniques this breakthrough SiteClick technology allows for antibody conjugation with complete confidence that the label will not directly interfere with the antibody binding domain. | antibodies, antibody labeling, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, fluorescent dyes, Qdot, western detection |
| Webinar | Fixed cell imaging—Five steps for publication-quality image With over 40 years dedicated to cell imaging research, we offer long-proven tools and protocols to help confidently create quality cell images the first time. This on demand webinar covers the 5 essential steps to getting great images. | antibodies, blocking, Celleste, dyes, EVOS FL Auto 2.0, fixation, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF), sample detection, sample labeling, sample preparation, signal amplification |
| Webinar | Learn to choose the right fluorophore when designing experiments The choice of fluorophore is one of the first important decisions to make in developing an experiment. Fluorophores are compounds that emit light at a specific wavelength when they have been excited at another, lower wavelength. Join our webinar and explore: How to choose the best organic dye for an assay Quantum dots and how they compare to other dyes When to use a phycobiliprotein like R-PE or APC When to use fluorescent proteins like GFP How to choose a suitable dye to match your instrument In addition, we will explore the basic characteristics, strengths, and weaknesses of the various fluorophores to help you choose and develop the best assay for your needs. | antibodies, antibody labeling, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, fluorescent dyes, fluorescent proteins |
| Webinar | A practical approach to antibody labeling The growing number of fluorophores available makes labeling your own antibodies a tempting proposition. But with many antibody labeling solutions available, selecting the best option can be a daunting task. In this webinar we will: Provide an overview of our antibody labeling kits Offer guidance on which methods are ideal for specific applications and experiments Provide tips and tricks to optimize your labeling protocol | antibodies, antibody labeling, flow cytometer/flow cytometry, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF) |
| Webinar | An introduction to immunofluorescence staining of cultured cells In this webinar, we discuss the steps of an immunofluorescent staining protocol including material list, common variations, and necessary controls. We'll also provide a simple troubleshooting guide and examine how to avoid common pitfalls. Presented by Jason Kilgore, Technical Support Specialist, Thermo Fisher Scientific. | antibodies, antibody labeling,antifades, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunofluorescence (IF) |
| Webinar | Basic techniques in autophagy research This webinar will introduce you to a series of analytical tools and techniques to help you identify and interrogate key features of autophagy. Topics to be covered include: Tips and tricks for selecting the right tools and achieving the best results Fluorescent proteins and antibodies used to analyze both live and fixed cells Analysis steps using a variety of multiplexing options, with quantitative methods for image analysis or fluorescence intensity measurement | autophagy, cell health, fluorescence microscopy/fluorescence imaging, fluorescent proteins, high content analysis, immunofluorescence (IF) |
| Webinar | Basics of multicolorflow cytometry panel design With the proliferation of new fluorescent dyes, as well as instruments that can detect 18 or more parameters multicolor flow cytometry has become more popular and more accessible than ever. This webinar presented by Dr. Holden T. Maecker at Stanford University will discuss the caveats of good panel design, including: Rules for designing panels Examples and practical application of these rules Controls and standardization Relevance of panel design to new mass cytometry platforms | antibodies, compensation, flow cytometer/flow cytometry, immunophenotyping, multicolor flow cytometry, viability |
| Webinar | A comparison of basic immunofluorescent labeling strategies In this free webinar, we will compare different immunofluorescent labeling strategies exploring the pros and cons of each method. You will learn when the use of a direct conjugate is appropriate and when amplification techniques can be utilized. We'll also present a simple decision tree to aid in determining the best method for each situation. | Alexa Fluor, antibodies, antibody labeling, fixed-cell imaging, fluorescence microscopy/fluorescence imaging, immunocytochemistry (ICC), immunofluorescence (IF) |
Not for resale. Super Bright Polymer Dyes are sold under license from Becton, Dickinson and Company.
For Research Use Only. Not for use in diagnostic procedures.
