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The gap year I intended to take between my Master’s degree and hypothetical Ph.D. is now going into its 4th year. Here’s why I’m not worried. These days it seems like undergraduates are proceeding en masse to graduate programs shortly after completing their senior year of college. An abundance of undergraduate research opportunities and poor…
One of the key characteristics of cytotoxic cells (i.e. CD8+ T cells, natural killer cells) is the presence of pre-formed cytoplasmic lysosomal granules. These structures house perforin and granzyme; two molecules that are essential for the lysis of target cells. Upon effector cell activation, granules are polarized toward the target cell and the contents are…
We have already looked into the different types of viral expression systems and when you might use one over another in my previous article. But why would you use viral transduction over similar techniques like plasmids? Just a reminder: Transfection is a lab technique where nucleic acids or proteins may be introduced into cells. When…
The sub-G1 assay for measuring apoptosis is easy, rapid, reliable, reproducible, and cheap and is widely used. However, you need to understand the mechanics of the assay and the apoptotic process, otherwise you could over- or underestimate your apoptosis results!
In part one, I discussed the ‘how to’ of simply freezing samples and the basics of vacuum evaporation, often referred to as speed vacing. Now, we’ll have a look at two more complex sample storage techniques (at least in terms of equipment) for drying samples (lyophilizaton and rotary evaporation) and the simpler method of blow…
IMS is a rapidly advancing technique that nicely blends the best bits of mass spectrometry with microscopic imaging. Learn more about it and how it could be applied to your work.
Generating a DNA vector library of single and/or compound mutants for a target protein can be a daunting task. If you’re lucky and work in a well-funded lab, you might outsource this process via gene synthesis. Most of us though, need to do it the old-fashioned way. Traditional QuikChange™ Traditionally, there are many steps you…
I’ve recently been doing some lyophilization of biological extracts. While I was preparing for the experiment, I became interested in the number of different methods there are for drying, concentrating and storing samples: freezing, freeze-drying, rotary evaporation, centrifugal evaporation and blow down drying. Here is a brief description of each technique for biological sample storage…
A PhD, like any worthy endeavor in life, requires hard work, patience and perseverance. It also involves challenges that will make you doubt your choice to pursuit an advanced degree in the first place. It is inevitable that a series of failed experiments or that one paper rejection will bring you down and temporarily make…
So you have a gene or protein that you think may be involved in migration or invasion and the next step is to embark on migration assays. These assays are useful for testing fundamental migratory processes, such as embryonic development, immune response, metastasis and angiogenesis. For a long time these have been an invaluable mainstay…
When choosing a model system for culturing tumor cells, we often think of the obvious tried and tested models. In vitro methods include cell lines that have been specifically selected to grow in culture flasks in incubators. While conveniently available, consistent and reproducible, cell lines are limited in that they may not represent the desired…
Finishing my third year of grad school, I can’t help but notice what a balancing act it has been- research and relaxation, finances and fun, beer and…not beer? I wanted to write this article to advise incoming first years and veteran grad students – even those mysterious 7th year students- on graduate school and the…
Flow Cytometry is a great way of seeing how many of your cells express a particular marker and how much of it is there. We do this by measuring fluorescence, but, as with all measuring systems, there will be signal that we are always trying to measure the above the noise. The signal that we…
Do you know about algae and their potential in today’s world? Do you know how to work with algae? Algae are becoming increasingly important in the research world.
One of the much sought after question asked by many researchers worldwide is – “What is the gene expression profile of a single cell within a heterogenous pool of cells?” While mass cytometry is the current ‘hot’ methodology for single cell analysis, the good old flow cytometry can help us perform rapid analysis of single…
gif by Kronin Okay not talking about that kind of matrix. But the kind of matrix that is crucial for MALDI (Matrix-Assisted Laser Desorption Ionization). I talked briefly about MALDI in my last article, Imaging Mass Spectrometry: The New(ish) Kid on the Block. Now I want to tell you more about MALDI, specifically the matrix….
From one lowly PhD student to another: we need post doctoral scientists. From their ability to seemingly do everything right to their moral support after a weekend of failed experiments and questioning the decisions that led you up to this moment * clears throat *. The list of support post docs offer is endless. So…
If you’re planning on using lentivirus for your next experiment, chances are you’re wondering how much virus to use. For in vitro work, multiplicity of infection (MOI) is the theoretical number of virus particles applied per target cell. That is to say, if you have 1 million cells and you want an MOI of 5,…
After ten years of postdoctoral research there is one important piece of advice I would give to anyone embarking on a research career: Spend as much time managing your data as you do generating it Take time at the beginning of each project to organize how you will record what you are doing day-to-day. The…
Three dimensional cell culture mimics the extracellular matrix (ECM) that offers the structure and support for cells in vivo, thus creating the complex architecture and network required for cellular communication. For 3D cell culture beginners (or enthusiasts), the information available may seem overwhelming. It sure was for me. But it can be simplified. For example,…
Human bronchial epithelial cells (HBECs) are a challenge to culture. As highly specialised cells that exist in carefully ordered multi-layered structures, they are especially fickle and finding optimum conditions to keep them happy is tricky. The cultures are also extremely sensitive to tiny changes in routine or environment. However, there are certain basic principles that…
If you are regularly doing ChIP-qPCR, ChIP-RNAseq or luciferase reporter assays to measure protein-DNA interactions, then this article is for you! ChIP experiments can tell you what DNA sequences your protein binds, and luciferase reporter assays predict whether your protein functionally binds a specific promoter to activate transcription – but a yeast one-hybrid (Y1H) assay…
I’m sure many of us are aware that the world of cancer research is exploding with the idea of cancer stem cells. This exciting hypothesis suggests that there is a small population of cells in the tumor that have stem cell-like properties. These stem-like cells are able to proliferate and differentiate into all the different…
A cover letter is a letter written to accompany a job application, a grant application, project reports, manuscripts, etc. that explains the purpose of your writing and incorporates certain key points of your application. It plays a decisive role in accomplishing the task it’s been written for, which is why it is so important to…
So it’s time to write up your thesis. By now, you probably consider yourself an expert on your topic or you hate the sight of it (or both!). Either way, hundreds of typewritten pages are all that stands between you and graduation/freedom. One of your first requirements will be to review the current knowledge on…
As a protein biochemist where bacteria were mere workhorses, imagine my surprise when I began work in a bonafide micro lab! I discovered that bacteria could be much fussier than my good ol’ cloning and expression friends E. coli DH5ɑ or BL21. One broth would not do for all, some even required blood! No, no,…
PhDs have been known for their nightmarish effects on students’ psychological wellbeing, to the point that the acronym PhD has also been dubbed ‘Permanent Head Damage’, ‘Philosophically Disturbed’ or ‘Please Help. Desperate’. Doing a PhD is an emotionally exhausting experience rather than being physically challenging. Here are some tips on how to survive the PhD…
As a dual-degree MD/PhD student I spent two years in the medical school doing classroom learning, now I’m in the lab trying to get a PhD, and then in a few years I’ll return for the last two years of medical school and work in the hospital. In this process, I’ve heard a lot about…
Lab work, as we are all aware, comes with many pressures: one of which is productivity. You want to generate as much quality data as possible to meet publication deadlines or perhaps the elusive thesis. Sometimes it may feel like hours spent in the lab don’t match the amount of data produced: for some this…
While it is true that there are some useful websites like SNPedia, or NCBI that can help you find rs codes for genetic variants, sometimes you need that info coming straight from the oven – particularly when you want to look at atypic SNPs or substitutions that have not been validated. So, in this post I…
Imagine trying to build a house without power tools: It’s completely doable – after all, people did it that way for centuries – but it’ll take you a lot longer and has limits. Similar to this, modern day biology now has its own set of “power tools.” So while you could do biology the old-fashioned…
Flow cytometry is a fluorescence-based technology, as is fluorescence microscopy and confocal microscopy. Fluorescence is fundamental to how a cytometer gathers data, but I am often surprised, as a core manager, at how little new users know about the process of fluorescence. So, this is where I always start the training process. Let’s get physical…
Western blotting uses electrophoresis and antibody-epitope affinity to give a semi-quantitative and (theoretically) clear measure of protein abundance. It’s a long procedure, filled with many steps—and even more room for error. Learning to troubleshoot certain problems is incredibly important for continued success with this technique. So what do you do when your final imaged product…
Take a look at the dotplot below, are you happy with the way it’s presented? Do you think that you could recreate that experiment? If you were a reviewer, would you accept that figure? Sure, it’s flow plot, it shows 3 populations of which two are gated. Read many journals and you will see data…
The word ‘retrovirus’ evokes images of HIV, the AIDS pandemic and a desperate worldwide effort to defeat this mighty adversary. But on the flip side, scientific research has managed to tame the virus and use it as a tool for advancement. Retroviruses are commonly used to introduce genes into mammalian cells to express or knockdown…
Genomic Science has come a long way since the early days of Sanger sequencing in the 1970’s. Today, there are jazzy new sequencing technologies that include fragment analysis, epigenetic sequencing, RNA/transcriptome sequencing and Next Generation Sequencing (NGS). Increasingly these technologies are becoming more accessible, but they still require highly specialized (read: expensive) equipment. Unless your…
As a scientist myself, I have always been focused on performing experiments to achieve my sole goal of producing data for a publication. Recently I started paying attention to the waste I solely generate on a regular basis in the lab. Starting from disposable Eppendorf tubes and pipette tips, to labels and Petri plates, it…
As a research intern this summer, part of my project included expressing and purifying a few proteins of interest. Two out of the three proteins posed no problem, but the third caused me to spend an agonizingly long amount of time– setting up new secondary cultures everyday, waiting for them to grow, forgetting to induce…
Extracting protein from tissue samples and cultured cells is Step #1 in many biochemical and analytical techniques. Before you can do a Polyacrylamide Gel Electrophoresis (PAGE), a Western blot, or mass spectrometry you need to extract your protein. Nowadays, a lot of labs have switched to kits for their protein extraction but these kits can…
Whether you’re just beginning, or nearing the end of your undergraduate career, getting involved in research can be overwhelming and stressful. There are many factors to consider, so keep it simple and focus on the most important ones: What area of research motivates you? What organisms do you find interesting? Are there any research professors…
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