Tissue Clearing from a Microscope Enthusiast’s Perspective


Posted by Ginny B. on Sep 11, 2019 3:15:00 AM

Biological imaging data has massive potential in terms of complexity of experiment and richness of dataset. While early imaging experiments were mostly descriptive, showing if "target X presents in cell of interest;" modern experiments have the potential to massively multiplex, define complex spatial relationships, measure levels or numbers of tiny subcellular components, and even measure mRNA.

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Topics: IF-IC, Neuroscience

Fluorescent Staining Using Multiple Antibodies


Posted by Tara W on Sep 13, 2017 3:00:00 AM


The use of multiple antibodies in a single experiment can provide useful information to researchers. Co-staining with multiple antibodies and cellular dyes is a simple, low-content form of multiplex analysis. Techniques for performing multiplex analyses in cells and tissues are powerful research tools that are applicable to general cell biology studies as well as diagnostic purposes. These techniques allow researchers to detect multiple biomarkers to assess their samples. They also allow for easy colocalization studies to determine relationships between analytes. Here we describe two common techniques for fluorescent staining using multiple antibodies in the same assay.

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Topics: Protocols, Antibody Performance, Primary Antibodies, IF-IC, Companion Reagents, techniques

5 Steps to Publication-Worthy IF Images


Posted by Tara W on Sep 6, 2017 3:30:00 PM

A picture is worth a thousand words, or in the case of immunofluorescent imaging, a thousand proteins. The images used to illustrate a scientific experiment should convey as much information as the text itself. Here at CST, we pride ourselves in the quality of our antibodies and our rigorous validation process. When we approve our primary antibodies for IF, we like to showcase them using high quality images generated in-house. Beyond our recommended IF protocols (check it out here), here are some additional considerations to make when planning your IF staining. 

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Topics: Protocols, Antibody Performance, Antibody Validation, IF-IC, techniques

Successful Immunofluorescence: Antibody Dilution and Incubation Conditions


Posted by Ken B on May 10, 2017 3:01:00 AM

Part four of a series on immunofluorescence techniques. Check out previous posts on Validation, Experimental Controls, and Fixation/Permabilization

After your samples have been prepared, it's time to incubate them with a well-validated antibody,  the workhorse of immunofluorescence. If you are a seasoned pro at IF experiments, you are probably used to checking the antibody datasheet (or web page) for the recommended dilution. But have you ever wondered where those recommendations come from?
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Topics: Protocols, Antibody Performance, Primary Antibodies, IF-IC, techniques

Successful Immunofluorescence: Fixation and Permeabilization


Posted by Ken B on Mar 15, 2017 3:00:00 AM

Part Three of a four-part series on Immunofluorescence. Check out The Importance of Validation and Experimental Controls.

The performance of an antibody is a crucial determinant in getting reliable immunofluorescence (IF) results. Equally important is the preparation of the biological sample - cells or tissue used in your experiments - before any antibodies are introduced. The fixation and permeabilization of your samples are key steps that can determine your experiment’s failure or success. The ideal fixative preserves a “life-like” snapshot while quickly stopping the degradative process of autolysis by crosslinking and inhibiting endogenous enzymes. This post provides examples of how different antibodies perform at their best using different protocols.

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Topics: Protocols, Antibody Performance, IF-IC, techniques, Fixation

Successful Immunofluorescence: Experimental Controls


Posted by Ken B on Mar 8, 2017 3:00:00 AM

 Part two of a four-part series on Immunofluorescence. Check out our posts on Validation and Fixation and Permeabilization

After months of hard work, your research has honed in on a hypothesis you can test with immunofluorescence (IF). You've chosen antibodies and performed pilot IF experiments (see The Importance of Validation), and the localization of the protein appears reasonable. But how can you be sure the IF data you've acquired represents real biological phenomena? We present two examples of experimental controls in this post.

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Topics: Protocols, Antibody Performance, Primary Antibodies, Antibody Validation, IF-IC, Post Translational Modification, Reproducibility

Successful Immunofluorescence: The Importance of Validation


Posted by Ken B on Mar 1, 2017 3:00:00 AM

Part one of a four-part series on Immunofluorescence. Check out Experimental Controls and Fixation and Permeabilzation.

After months of hard work, your research has zeroed in on a hypothesis you can test with immunofluorescence (IF). But now you have to make a choice. How do you decide which antibody to use to get reliable IF results? How do you know if the images are accurately reporting the target's localization? We explore some considerations in this post.

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Topics: Protocols, Antibody Performance, Primary Antibodies, Antibody Validation, IF-IC, Reproducility

3 Questions to ask before starting your next immunofluorescence experiment.


Posted by Liana G on Oct 19, 2016 3:00:00 AM

Specificity, consistency, and optimized assay conditions are three key elements that help ensure reliable immunofluorescence (IF) staining results each and every time. So, before starting an experiment you should ask yourself the following three questions:

  • Is your antibody specific?
  • Is your antibody supported by an optimized IF protocol?
  • Is your antibody performing consistently?

Let's take them each in turn...

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Topics: Protocols, Primary Antibodies, Antibody Validation, IF-IC, Cell Biology

3 Questions to ask before starting your next immunofluorescence experiment.


Posted by Carolyn P on Sep 7, 2016 3:00:00 AM

Specificity, consistency, and optimized assay conditions are three key elements that help ensure reliable immunofluorescence (IF) staining results each and every time. So, before starting an experiment you should ask yourself the following three questions:

  • Is your antibody specific?
  • Is your antibody supported by an optimized IF protocol?
  • Is your antibody performing consistently?

Let's take them each in turn...

READ MORE >

Topics: Protocols, Primary Antibodies, Antibody Validation, IF-IC, Cell Biology, techniques

5 Steps to Publication-Worthy IF Images


Posted by Tara W on Sep 23, 2015 8:00:00 AM

A picture is worth a thousand words, or in the case of immunofluorescent imaging, a thousand proteins. The images used to illustrate a scientific experiment should convey as much information as the text itself. Here at CST, we pride ourselves in the quality of our antibodies and our rigorous validation process. When we approve our primary antibodies for IF, we like to showcase them using high quality images generated in-house. Beyond our recommended IF protocols (check it out here), here are some additional considerations to make when planning your IF staining. 

READ MORE >

Topics: Protocols, Antibody Performance, Antibody Validation, IF-IC

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