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You are probably familiar with fluorescent in situ hybridization (FISH) to detect and localize the presence or absence of specific DNA sequences on chromosomes. But did you know there are numerous FISH experiment variations? Including high-resolution FISH and quantitative FISH? Read here about Fiber-FISH, Q-FISH, and Flow-FISH and decide if you would like to undertake one of…
If you are cooking up a way to test if two proteins interact, you need a yeast two-hybrid (Y2H) screen in your recipe book. You don’t want your Y2H to turn out half-baked – so check out this guide to Y2H and we’ll help you make sure yours will rise to the occasion! What is…
It is incredibly important for aspiring young scientists to keep up to date with the scientific literature. We all know that some journal articles are a slog, and critiquing other’s research is often an onerous task. Sometimes it’s good to have a break. What follows is a list of popular science books I have found…
If you are establishing a flow core facility or even setting up just one cytometer, then you need to know a few things before you plan your new lab and start looking at purchasing cytometers: Before You Start 1. Know Your Customer It’s really important to know if you are going to be setting up and…
Want to measure how much mRNA you have in a particular sample? Easy! Make some cDNA, add some fluorescent DNA-intercalating dye, pop it into a quantitative real-time PCR (qRT-PCR) machine and Bob’s your uncle! You have your result! Easy right…? Not so fast. As with any scientific assay, qRT-PCR requires some optimization. First, you need…
Would you eat your spaghetti dinner without a plate? No, of course not! It would make a big mess and be ugly to look at. Instead you NEED something to put that spaghetti on, to contain it, to keep it clean, to make it look nice – a bowl, a plate, SOMETHING! By the same…
“You think you know, but you don’t know and you never will, okay?” was the response an irate Jim Mora, head coach of the New Orleans Saints, gave to an unwitting journalist questioning his management – his point being that unless you’ve actually been in a professional sports team, you will never know what it’s…
The NIH grant that you are working on only gives you five pages to describe your research strategy. You’re wrestling with a research report for Science that has a maximum word limit of 2500. And the abstract for the conference you’d like to speak at this spring only allows you 300 words to summarize the…
It was not long since the commercialization of NGS (a little more than ten years ago) that scientists went beyond the basics and got creative with the new technology to study much more than just the sequence of DNA. In this article we highlight some of the different NGS technologies and methods available out there….
Science is an expensive business and those who use high energy-demanding techniques may not even realize just how expensive they are. The Cost of PCR Let’s looks at PCR. You need to pay for the machine, all the ingredients including expensive enzymes, a freezer and a fridge for your ingredients, tubes and caps, not to…
Dear Aunt Yersinia, A very annoying postdoc in our group keeps telling me off for spinning E.coli at 13K in a tabletop centrifuge. The postdoc claims that high speed damages cytoskeleton and this will reduce my transformation frequency. But I don’t believe her as the cells are cushioned by water during centrifugation. Can you tell…
No matter how we make measurements, there will be variation (a spread of data). Take 100 people and ask them to guess your age and you will get a range of results: some will be too low (excellent!), some too high (not so good!). It is the same with any of our laboratory experiments –…
Plasmid incompatibility could ruin your experiments before you’ve even started. And if you don’t know what it is, you can’t troubleshoot it! This article explains plasmid incompatibility, so you can’t be caught out by it.
Detergents are all around us in the lab – and that’s a good thing! Thanks to their chemical structure, detergents can solubilize and interact with many types of molecules, making them vital to research. To show you why detergents are such a good thing for scientists, we’ll go through six examples in molecular biology where…
Labs across the world spend a great deal of money on Taq polymerase. Find out how to save your lab some money.
Whether you are writing your thesis, a manuscript or a grant, there will come a time when you need to write, but getting words onto the paper will be like trying to get DNA from a rock – you are pretty sure it ain’t going to happen. Luckily there are a few tricks you can…
“Making the simple complicated is commonplace; making the complicated simple, awesomely simple, that’s creativity.” – Charles Mingus Next Generation Sequencing (NGS) technology has boomed in recent years, allowing researchers to probe further into the workings of the genome. According to the theory of simplicity, it is the simple principles on its basis that make…
Problems with expressing your gene? One of the potential stumbling blocks in heterologous gene expression is incompatible codon usage. Every amino acid can be encoded by more than one codon, and for every amino acid each organism has a favorite codon that it tends to use more often than the others. The availability of tRNA…
mRNA, miRNA, siRNA, tRNA, rRNA! Just what do all these RNAs do? Most biology graduates will have heard a good deal about mRNAs, tRNAs and rRNAs since these are vital players in protein synthesis. For siRNAs there has been a lot of focus within drug discovery and biomedical research over the last decade, but there…
If you’ve ever performed PCR, you’re probably already very familiar with DNA oligonucleotides (or oligos). But did you know that these molecules can do so much more than just act as simple primers? You can add a wide range of modifications to your oligos, which may change the stability, binding, solubility and even visibility, to…
Discover the different approaches to FRET quantification and get tips for selecting your FRET pairs.
Do you need your protein in its native form, intact, with full functionality? Do you need to isolate organelles and nuclear fractions from the cytoplasm? Or do you need a slurry of everything in your cell or tissue? Whatever your experiment, you can maximize the amount of functional, detectable or active proteins by handling your…
Once you have been in the field of Flow Cytometry for a bit, the name Howard Shapiro will be familiar to you. However, for those who are new to Cytometry, you might not be aware of Dr Shapiro and his fabulous book ‘Practical Flow Cytometry’. Most Cytometrists have a paper copy of this bible of Flow…
With in vitro translation (IVT) you can produce protein in a tube. Cool sure, but would IVT be a good choice for your experiments? Not sure? Then read on. Here I will cover the strength and weaknesses of IVT. How In Vitro Translation works First things first, for IVT you need ribosomes. While theoretically possible,…
There are about as many protocols to prepare coverslips as there are ways to make tuna casserole. You can spend from 5 seconds to 2 days, depending on what your lab prefers. But in the end, what’s really needed? I’ve tried many protocols over the years and I’ve questioned some steps. Here I share my…
I previously wrote an article for BsB detailing my experience transitioning from lab-bench research into research administration roles after a particularly unhappy experience as a postdoc. About a year into my second research admin role some restructuring occurred and I decided to try to move back into the lab. I am now working again as…
If you want to get the maximum yield and quality from your next-generation sequencing experiment then you are going to need to make sure each of the libraries you produce is carefully quantified ready for pooling and/or loading onto a flow cell. If the quantification goes wrong you’ll get a bad balance of samples within…
After designing a multicolor flow cytometry panel and securing the necessary cells and reagents, the process of optimization of the panel can begin. The first step in that optimization is titration of your antibodies. In this process, following a standard protocol to be used in the final analysis, you stain a known amount of cells with…
One of the really exciting parts of being involved in research is the opportunity to travel to a conference (hopefully at an exotic location!) to present your work and get to see presentations from the major players in your chosen field. Now you’ll probably have all sorts of frivolous reasons for not wanting to go…
Loop-mediated Isothermal amplification (LAMP), is an emerging technology that allows DNA amplification at a constant temperature. The key to this principle is the use of a DNA polymerase that possesses strand displacement activity. As a result of this property there is no need for heat denaturation of double stranded DNA in order to allow primer…
One of the most annoying traits of “classical cloning” is an imperfect system of discriminating between the clones containing an empty vector and vector with insert after cloning. Even when your self-ligation control plate is empty, you can have a lot of colonies containing an empty vector on the “vector + insert” plate. Even the blue-white…
There are so many unspoken rules to working in a lab! It’s unnerving what will cause frayed nerves to snap, people not to trust you and a good relationship to turn sour. Here are some of the rules I’ve learned. Feel free to add more in the comments section below. 1. Thou Shalt Not Touch…
In my previous article ‘Choosing a scripting language for next gen sequencing: Python, Perl, and more’ I discussed several of the more common programming languages used for next generation sequencing and things to consider when picking which one to learn. But now that you know WHAT you want to learn, HOW do you go about…
You use your antibody frequently, maybe even every day. You rely on it for western blotting, immunohistochemistry, FACS, ELISA, and immunoprecipitation. You’d be lost without it. But how well do you really know your antibody? Are you sure that it detects what you believe it to detect? If you have the slightest doubt, or if you have…
Every protocol for single cell PCR can be broken down into two steps. In the first step, the cells are isolated by micromanipulation, laser capture microdissection, flow cytometry, or by direct micropipetting. Next, the genetic material is processed by PCR to amplify your sequence of interest. Here, we’ll go through the different options for isolating…
When it comes to publishing your paper you want to show the world what excellent research you’ve worked so hard to produce. Part of that is providing enough detail so that others can reproduce your work and take it further. There are certain details you need to include in your paper; many, but not all…
“The greater the power, the more dangerous the abuse.” – Edmund Burke “With great power comes great responsibility.” – Uncle Ben (quoting Voltaire) While it’s unlikely that either of these speakers performed multi-dimensional flow cytometry, it is important to remember these quotes in the context of developing and implementing a good polychromatic flow panel. More fluorochromes are…
One of the best parts of R is how extensible it is. Over the years, the community has put together hundreds (thousands?) of amazing packages to make your workflow easier. The downside of this wealth is that it can be hard to find packages that do exactly what you want! Therefore, I’ve put together a…
As with any experiment, choosing the right personal protective equipment is essential. In this series we’ll take a look at what different types of hoods add to your arsenal of PPE, what they do and how you can benefit by using them. First, why do I even need a hood? It’s common to run a…
It’s a familiar story – molecular biology meets protein science, they get closer and sparks fly. But how exactly does a proximity ligation assay (PLA) work and how do you make sure yours will have a happy ending? What PLA Does PLA allows you to detect individual proteins or individual protein interactor pairs without ectopic…
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