Search results for: western blot
The 4 Important Steps for Western Blot Quantification
…blot quantification. Why Should You Perform Western Blot Quantification? First, what do we actually mean by western blot quantification? Put simply, this refers to the measurement of the signal emitted by…
Read MoreThe Essential Guide to In-Cell Westerns
…Western blots, and ELISAs. Property In-Cell Western Traditional Western Blot ELISA Target Protein targets are inside fixed, whole cells. Targets are purified from cell lysates or biological matrixes. Targets are…
Read More3 Hot Tips to Optimize Your Western Blot Transfers
Does your western blot look like someone sat on black playdough? If you’re lamenting over yet another blotchy or bad western blot, don’t fret. The cause can often be a…
Read MoreFluorescent Western Blotting: Lowdown and Advantages
…Therefore, to ensure reproducibility and accuracy of your western blot results, IR fluorescent western blot is the way to go. 5. Extended Signal Duration Fluorescent signals are extremely stable. You…
Read MoreKnow Your Western Blot Jargon! A Quick Review.
…quick review of SDS-PAGE (Polyacrylamide Gel Electrophoresis) Western blotting including its required jargon (see bolded and hyperlinked words). The Purpose of a Western Blot The goal of a Western blot…
Read MoreWhat Ponceau S Staining Can Reveal About Your Western Blot
…of Ponceau S staining of a western blot membrane. (Image Credit: Thomas Warwick.) Three key benefits of Ponceau S stain to note: It does not affect the protein structure. It…
Read MoreStreamline Your Western Blots
…western blot by eliminating the second antibody incubation along with the washes between the two antibody incubations. I have used this method for around five years, and have yet to…
Read More5 Ways to Destroy Your Western Blot
…that are easy to mess up. How do you make sure that your Western blot is successful? Avoid the following five ways to destroy your Western blot: [Edit: Now this…
Read MoreHow to Choose Quality Antibodies for Successful Western Blotting
…Want all your Western blot buffer recipes, an ECL reagent recipe, and an easy protocol all in one place and on hand? Download Bitesize Bio’s free Western blot cheat sheet….
Read MoreSecond Chance Saloon: How to Western Blot a Coomassie-stained gel
…tested this hypothesis recently using a gel that had been freshly stained with InstantBlue. I increased a wet-blot transfer time 1.5 times, but otherwise followed the usual Western blot protocol…
Read MoreHow Do YOU Image Your Western Blots?
The last step in western blotting is imaging the blot – this is the moment of truth, when you finally get to see the results of the experiment you’ve been…
Read MoreGet your stripping stripes! Find out how to strip and re-blot your Western
…antibody itself, so dissociation of your antibodies won’t remove the coloured bands. Keep your blot wet. Your blot can’t be allowed to dry out at any point after transfer as…
Read MoreHow To Make ECL Reagent: 4 Ingredients, Better Blots
What’s more disappointing than a failed Western blot? Several days of work and effort—gone. After the transfer step, the second biggest cause of failed Western blots is the ECL reagent….
Read MoreThe Top 10 Western Blotting Mistakes (and Solutions!)
…blots! Want all your Western blot buffer recipes, an ECL reagent recipe, and an easy protocol all in one place and on hand? Download our free Western blot cheat sheet….
Read MoreBlock, Stock and Barrel – A Guide to Choosing Your Blocking Buffer
…signal on your final Western blot. Yep, you guessed it…I’m talking about blocking. The membranes used for Western blots have a high affinity for proteins, which is a good thing…
Read More3 Approaches to Western Blot Transfer
…Agrisera. Want all your Western blot buffer recipes, an ECL reagent recipe, and an easy protocol all in one place and on hand? Download our free Western blot cheat sheet….
Read MoreNon-specific Binding? Tips to Sharpen up Your Western Blot
…through some ways to sharpen up your blot, in order of relative importance. Blocking This is the most important step of the blot—if you don’t block the unoccupied sites on…
Read MoreHow Good Is Your Best Friend? A.K.A. Have You Validated Your Antibody?
…that their antibodies work via a western blot. Such antibody validation efforts typically show a band at the expected molecular weight, but nothing more. These western blots do NOT tell…
Read MoreGet Out of Western Blot Hell: An Intro to Mass Spectrometry
…(and slow!) way by running a multitude of Western blot. But rarely do labs have unlimited funds for Western blot antibodies. And lets face it, do you really want to…
Read MoreTen Tips for Turning Beastly Western Blots Beautiful
…that could turn a really ugly Western blot into a thing of beauty . To save you the humiliation of presenting an ugly blot at lab meeting, I am sharing…
Read MoreStripping Blots – It’s All Fun and Games Until Someone Loses Their Protein
…say you run a blot for protein X and it provides a signal of “10” on your blot. Then you strip, re-probe, and run a second blot for protein Y,…
Read MoreHow to Optimize Your Western Blot Transfers
…your Western blot. Transfer times are empirical and based on your own particular samples, which means that there is no easy way to determine how long you need to run…
Read MoreWestern Blot, ELISA, SPR, Biosensor Assay or PCR: Which Technique Should I Use?
…right method according to your specific needs. When Should You Use a Western Blot? Standard western blotting can detect the activation of proteins involved in a signaling pathway. Activation of…
Read MoreAnalyzing Cell Signaling with Flow Cytometry: Go with the Flow
…that’s available for phosphorylation. Today, I am going to tell you why you should put your old trustworthy western blot aside and give multi-parameter flow cytometry a try when it…
Read MoreFilm vs. Digital: A Real Western (Blotting) Showdown
…darkness for your blot, you’ve managed to grope your startled colleague. Once the shrieking subsides, you may be able to locate the developer and develop your blot…but your colleague will…
Read MoreMore Than a Clever Name: Northern Blots
…big brother: The Southern blot. Southern blots are a method of detecting DNA and named after the surname of their inventor Ed Southern. A typical Southern blot experiment goes as…
Read MoreFishing for Kinases with Multiplex Inhibitor Bead Assays
…column, their method excludes the possibility of using Western blot. However, this isn’t a serious drawback because Western blot is notoriously unreliable, especially in the detection of low abundance proteins…
Read MoreConstant Current or Voltage in SDS-PAGE: The Great Debate
…Resistance (R) Feeling Hot, Hot, Hot! If you’ve ever performed PAGE or a western blot transfer, you may have noticed that the chamber can get pretty warm. I’ve even heard…
Read MoreSouthern (blot) exposure remains a useful technique
At a meeting recently, I asked two PhD molecular biologists about the last time they used a Southern blot. After nearly a minute of unrestrained laughter, they asked “Who on…
Read MoreFractionating Proteins by Solubility: or, How I Learned to Stop Worrying and Love the Ultracentrifuge
…in the sample and solubilize the proteins. Perform a Western Blot to Determine Whether Your Protein of Interest Is Soluble, Membrane-Associated, or Both When performing your western blot, remember to…
Read MoreAntibody Validation
…western blot depends on the denaturing of proteins. Consequently, a western blot-validated antibody may work fine in denaturing conditions but may fail to recognize antigens in their native conformation (i.e.,…
Read MoreSix Fixes For Antibody Co-Elution In Immunoprecipitations
…co-eluting. Try strategies 1, 2 and/or 3. If the co-eluted antibody obscures bands on a western blot: Then you don’t necessarily need to stop the antibody from co-eluting, you just…
Read More5 Easy Ways to Tell Salt and Protein Crystals Apart
…can make is spending months purifying the wrong protein. Why haven’t I written an article on this? Because it’s easy enough to avoid—do a Western blot on your purified sample…
Read MoreHow Do YOU Make Sure That Your Western Blots are Evenly Loaded?
For Western blot data to be reliable, it is important that you load known amounts of sample into each lane of the gel. This is of particular importance if you…
Read MoreQuantifying Individual Proteins Per Bacterial Cell
…Determine Linear Range of Detection for Western Blots When the protein is purified and the lysates are prepared, it’s time to set up western blots! First conduct a western blot…
Read MoreDetecting Post-Translational Modifications
…can point to the site of the PTM in the protein molecule. The technique can be used to more precisely analyse protein fragments separated by Western blot, IP, and other…
Read MoreHow To Preserve Your Samples In Western Blotting
When running a quantitative Western blot, it’s crucial that your sample preparation is consistent. Incomplete protein extraction from one sample will skew your results when you compare it to the…
Read MoreHow To Preserve Your Samples In Western Blotting
When running a quantitative Western blot, it’s crucial that your sample preparation is consistent. Incomplete protein extraction from one sample will skew your results when you compare it to the…
Read MoreFive Fast Tips for Blotting Large Proteins
…steps, you should have a beautiful blot showcasing your large molecular weight protein in no time! Figure 1.Western blot of a high molecular weight protein. Image courtesy of Victoria Brown….
Read MoreBuying a Secondary Antibody: Why all the Choices?
…are conjugated to fluorescent dyes and are used in FACS experiments, in immunofluorescent detection of proteins in cells or tissues and more recently, in Western blot analysis using CCD detection….
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