I think most people would be surprised at how many cover letters, resumes and back-and-forth emails are filled with incorrect spelling and grammar. This is 100% inexcusable and may cause you to lose a position before you were ever considered. Additionally, if you are committing these errors, you probably do not know it and have committed to same error many times over.

The solution - SPELL CHECK! Spell check everything, including your resume, cover letters, emails, LinkedIn profile, Facebook site, etc. After you spell check it, do it again for good measure. Copy and paste your information into multiple formats to check for errors. Ask peers and friends to review your documentation. Each of the documents you employ in your job search will be used over and over; the more refined they are, the better you will look to perspective employers.

The same goes for communicating with your recruiter – take the time to use proper spelling and grammar. Recruiters are much more likely to represent candidates who can communicate clearly and don’t make simple errors.  Every candidate a recruiter represents is a reflection of their company to their client.  Companies have passed on some of our candidates because of misspellings in a thank you note. Do not underestimate the crippling impact of poor spelling or grammar!

But beware, spell check and grammar check cannot and will not catch everything. Read, re-read (and even read it backwards). Ask others to review your information when appropriate. There are many aspects of your career search that you cannot control, but spelling and grammar are completely within your grasp. Control what you can – measure twice, cut once.

Now I would like to discuss some ways to be the lab favorite, also known as the “golden child”. Does your lab have a “golden child”? Someone who is always perfect, the favorite of the PI, the go-to person for everything by everyone? Do you wonder how they got that way?

Well, actually, I can’t tell you because I wasn’t the golden child of my lab.  But I’ve known a few and I run a lab myself. So instead, I am going to tell you what qualities or behaviors a person has to show to be my golden child. That’s right. Here is a list of 5 ways to be my #1 scientist. You’ll be on my “awesome” list instead of the s-list if you do the following:

1. Look things up yourself. Don’t go to your PI or to the post-docs with questions that are easily researchable yourself.  If your question is “should I do it this way or that way?” you should already have an answer as to what you think you should do. Your PI wants to see that you are learning how to design experiments and direct a project on your own.

2. Take initiative. If you have an idea or a theory, go for it! Try it without asking. Please do not ask me before doing every experiment. Just do it (within reason- if it requires a trip to Mexico to collect rare mud samples from ancient Aztec burial grounds, then of course, discuss that first) . I expect my scientists to think on their own and have ideas and it is ok to try them out first and see if they have value.

3. Show leadership. You are the project manager for your research. Getting your project done may mean collaborations with others both inside and outside your institution. This is a repeat of points 4 and 5 of the article “Pointers for new graduate students“.  In an industrial lab,  I like to see my scientists communicating with researchers working in areas of common interest and taking the lead in working together to generate data for posters or for future products. In academics, you’ll want to discuss with your PI first what labs you want to collaborate with, to protect the confidentiality of your work.  Once you both agree it is a good idea, take the lead in this area.

Showing leadership can also be willingness and ability to manage another person, such as an undergraduate or a rotation student, and doing a good job at it. Showing the ability to lead another person’s work, oversee someone else’s progress, and take responsibility for their results shows excellent leadership.

4. Be the expert in your area of research. You should know everything about your subject of study. Know what’s going on in the field and who the thought leaders are.  Your PI can’t read all the details of every paper for every project in the lab. They are counting on you to do that. Here are some suggestions on becoming an expert.

5. Show progress each week. We all have our off weeks (sometimes months- dare I say year?) where things just don’t go right. But as Nick mentioned in an earlier article, there’s no need to worry about getting results as long as you are doing things properly. If you can show logical experiments as to how you are approaching the problem and how you are dissecting it, it gives your PI confidence that you can figure it out.

If someone comes in with inconsistent results week after week and can’t figure out why and doesn’t have a plan for getting to the answer another way, they are going to quickly fall down the totem pole. If I have to hear “It didn’t work”, I want it to be followed by “but I think I know why or what to try next.”

6. Other stuff. Some things are obvious- of course you should be helping others when you can, sign up to make the coffee for journal club once in a while, bring the microbrewed beer for Friday happy hour (instead of the Pabst Blue Ribbon), and get the extra discounts for the lab by staying on good terms with the sales folks.

For a busy PI with way too many commitments and not enough time to read all of their emails, the best grad students and post docs are independent and inventive, in control and motivated to succeed. Even as a new student, it is never too late to show your willingness to think for yourself and think outside the box (in my lab, we call it “cowboying it”) .

The most fun we have in R&D at MO BIO is when we get to “cowboy it”. And usually our biggest leaps are made this way too.

I would love to hear from some of you “Golden Child” grad students. I know you are reading. Let’s go superstars. Add to this list and tell us more about how and why you are #1.

~Suzanne (now on Twitter)

So it makes total sense that for science, we would learn techniques via the internet. But not just a literary “how-to” guide anymore. Videos as a learning tool are starting to take a bigger role in teaching. A live demonstration speaks a thousand words!

In a previous article, I talked about JoVE (Journal of Visualized Experiments) and how they were leading the way in publishing live videos on techniques that go along with a published paper.

But learning can also be just a simple 30 second to 5 minute video on how to perform a specialized technique, one which is  not necessarily publishable or citable. That is what some scientists are doing from both academics and industry.

Below are two examples of the use of video to teach techniques; their sole purpose is to teach.

How to Handle a Water Filter Membrane

This first one was made by MO BIO Labs and demonstrates a best practice for transferring filter membranes from a vacuum filter unit into tubes for bead beating or storage. This short 50 second video is enough to give the scientist the basic technique required to safely handle a wet filter membrane without tearing or scratching it.

Filter membrane insertion into a bead tube

It is short and helpful- allowing the user to visualize a technique that in words would be difficult to emulate.

Making a Agarose Gel

This next video was made by researchers at the University of Leicester. Even though this is longer (5:44), it is packed with great information on agarose gels including the size range of agarose separation, what % gel to run for small and large fragments, and what your gel will look like if you don’t melt the agarose completely. This video would be perfect for training new students. This lab also did a second video called Running an Agarose Gel.

University of Leicester Making an Agarose Gel

As you can see, this video is more professionally done and the lab far cleaner than most I’ve ever been in. I hope the University of Leicester plans on doing more molecular biology videos!

BiotechprojectAZ Channel

I wanted to draw your attention to one more academic institution that is creating videos for learning and that is called BiotechprojectAZ. These series of videos show undergraduates demonstrating how to prepare, pour, and stain agarose gels. They are very nicely done and the students speaking on the videos did an excellent job.

These videos are all new and unfortunately, I can’t find any more detail about the location of this work- whether this is a university or college or if AZ stands for Arizona. If the director and producer of the BiotechprojectAZ channel would contact us, we would love to hear more about your plans for future videos and why you decided to embark on this great project.

As you can see, videos offer a better way of learning, but especially when it comes to science. Some much of science is technique based and having “good hands”. If the person with great lab hands could show us exactly how they do that method, the mystery would be revealed.

Bitesize Bio is interested in featuring your video techniques for our readers.  They don’t need to be professionally produced and they can be short or long. If you have a great way of doing some technique and want to share it with other scientists, send us the video and we will post it on Bitesize Bio.  Scientists helping other scientists is what we are all about.

But to scientists inexperienced in this technique, who typically see the results of IHC/ICC experiments in the form of pretty pictures, it can certainly seem like alchemy.

While these pictures can be pretty in their own right, the pictures display a range of important scientific information. The illusion of seeing such a pretty picture is the observer often thinks the process, time and skill involved in obtaining these images is relatively little and easy to do. Nothing could be further from the truth. A good portion of IHC/ICC experimental conditions is all determined empirically. Trial and “error” is part of all scientific research, and IHC/ICC is no exception.

The purpose of this article is to give the beginner a list of some initial variables one must think about before attempting his/her first set of IHC/ICC experiments. I have one caveat: there is perhaps an exception to most of I may say below. Remember, in research as in life, not everything is an absolute.

So here’s what you need to achieve gold (rather than lead) standard results from IHC/ICC…

1. Fixation. In real estate investing, a famous phrase which describes the potential investing success, is often “location, location, location”. The same can be said for IHC/ICC, “fixation, fixation, fixation”, this step is the first and perhaps the most important major step in obtaining successful results in IHC/ICC experiments. The wrong fixation protocol will almost guarantee you get undesirable results, that is, no signal from the protein you are trying to observe in your sample. Fixation protocols themselves have a wide range of variables, such as: the chemicals used to fix the sample (paraformaldehyde, methanol, acetone mixtures and others!), the temperature (4° C to 37° C) and time (15 min to overnight) for which a sample should be fixed.

2. Primary Antibody Selection. Will you use monoclonal or polyclonal antibodies, when should you use one over another? Who should you buy from? I’ll cover this in a future article, the answers can be lengthily!

3. Controls. There are two types of controls one should use, a negative control and a positive control. Each control tells you something different, and is used to help insure an accurate interpretation of your results.

A negative control, where you are using a primary antibody and a secondary antibody in the experimental group, consists of the omission of the primary antibody in the negative control. This negative control will give you an idea of how much signal, if any, is due to the secondary antibody alone.

Another type of negative control would substitute the primary antibody for non-immune serum or even better pre-immune serum, followed by detection with the secondary antibody.

A positive control tells you two things:whether your technique is sound, and whether the primary is detecting the appropriate target. Positive controls can come in many forms depending upon the type of experiment. In the case of IHC, a separate slide with tissue known to be immunopositive for your protein of interest should be used.

In the case of ICC, where the experimental group is cells transfected to express your protein for example, using parental cells should yield no specific signal due to the primary antibody.

You have to remember, if you are labeling your protein of interest with an antibody, you may observe background labeling. The background may be the result of binding by the primary, secondary or both. Antibodies are inherently “sticky” proteins by nature, so an antibody may not necessarily bind to only the proper epitope, thus generating a false positive. If you have the proper controls in place, these will only make your results more compelling should you get a true signal.

4. Signal Detection. In short, will you use a chromagenic substrate, which requires brightfield microscopy to observe the staining pattern, or will you use fluorophores commonly conjugated to antibodies to see the fluorescent signal? The answer depends on your needs.

Do you need to see your sample’s structure and only one protein, if so, then use brightfield. Or do you need to label two or more proteins for colocalization purposes? If so, then use a microscope equipped for fluorescence.

In the age of computers it is possible with the right equipment to overlay a fluorescent image onto a brightfield image, thus maintaining the ability see both cellular structure and protein localization. However, if you are only using a single label and require the sample’s structure, you will typically achieve better results by using brightfield microscopy.

5. Microscopy. With the recent advances in microscopy it is possible to use a variety of microscopy technologies to observe your results. If you have a core instrument facility, and you aren’t sure what to use even after reading and asking friends, I recommend speaking to the Director of the facility for advice.

Generally speaking, people get sharper images with laser scanning confocal microscopes than they do with compound microscopes. However, to be fair, this is dependent on your sample as well. I have heard from researchers who have used a compound microscope equipped for fluorescence getting better images from their samples than they do on a confocal microscope, so know your sample!

6. Image Processing. Many people use a variety of software for post-processing of images which are typically digitally acquired. One essential tip is to always maintain the original image file. Never make changes to the original image by saving over it, save the changes as a new file. Once you lose the original you are in trouble should you ever need to reproduce the original, especially to a scientific editorial board, or for a university/federal investigation.

In addition, before you make changes to the images, decide which journal(s) you are going to submit to and see what their criteria are for the acceptable altering of original images.

Some journal publishers now have forensic image analysis teams and all submitted images are put through special software which can identify what types of changes were made to the image. Scientists have been asked to reproduce the originals, or simply go back and alter the original image with only the accepted types of changes allowed by the journal.

IHC/ICC is a really useful technique but, like all useful techniques, only if it is carried out with due care and attention. I’ve outlined here some of the variables you need to consider before starting your experiments to ensure that your efforts bring in a pile of golden results.

Do you have any IHC/ICC tips or questions?

1. Save money by recycling electroporation cuvettes
Article: Re-cycling Electroporation Cuvettes

2. Ditch the LB and get rich cultures for super-productive plasmid preps by using PDM, a stoichiometrically optimised medium for plasmid production.
Article: Pimp Your Plasmid Growth Medium

3. Using LB-grown cultures for IPTG induced expression? Lactose in the LB will give a low level of background expression. Switch it off by adding 0.3% glucose.
Article: Shut of Background lac Promoter Expression in LB”>

4. Ever noticed that when you do the PE wash during gel extractions, a droplet of PE tends to stubbornly rest inside the column at the side that was to the outside of the centrifuge rotor? Get rid of that droplet, and stop it carrying through during elution, by turning the tube 180 degrees so the droplet is on the inside, and giving it another quick centrifugation.
Article: 5 More Tips for DNA Gel Extraction

5. Eppendorf tubes sent in standard envelopes can get stuck in mail sorting machines. So when you are sending plasmids in the mail either use a padded envelope, which doesn’t go through the sorting machine, or soak the plasmid in a small piece of filter paper, wrap it in cling film and send it flat packed!
Article: Sending Plasmids: How to Avoid Jail Time and Shredded Envelopes

6. Did you get too many bands in your PCR reaction? Here is a fast and easy method to rescue your PCR. Run the PCR reaction on a gel to separate the products and then stab the band of interest with a 20G syringe needle a few times. Swirl into a new PCR reaction and 20 cycles later, you’ve got your single band ready for cloning or sequencing.  This is fast, easy, and much less expensive than PCR optimization.
Article: PCR Rescue: Making One Band From Many

7. Another good way to eliminate non-specific PCR amplification is the Touch-Down PCR (TD-PCR) technique. TD-PCR allows for amplification of only the amplicons with the highest primer-template complementarity.
Article: Touchdown PCR: a primer and some tips

8.  Want to become the lab MacGyver?  Here is a top ten list of lab MacGyverisms; ways to use everyday items to make gadgets and low-tech solutions for the lab. Examples include using McDonald’s straws (they are autoclavable!) for disposable pipettes and cheap liquid handsoap for blot washes. Setting up a lab at home seems very feasible after reading this article!
Article: Low Tech Lab Gadgets: My All Time Favs

9. There are numerous competent E.coli strains to choose from. For clarity on the benefits of all the different mutations and then how to choose the right strain, this article outlines the options simply and clearly.
Article: Choosing a Competent E.coli Strain

10. TBE sucks. Run gels for 10 minutes at 200 volts using sodium boric acid buffer instead. 10 mM Sodium Boric acid stays cool even at high voltage, allowing gel gels to run 7 times faster!
Article: Faster, Cooler, DNA Gels

Have any more tricks of your own? Share them with us in the comments, or using the contact form. If you’ve got a good one we’ll write about it — or you are always welcome to contribute an article yourself.

Happy Halloween!

Well Bioconference Live allows you to do just that on 17-19 November (2009). And I think this is a not-to-be missed event. As well as lining up some great speakers, the Bioconference Live team have worked hard to capture the conference experience, offering the opportunity to field questions to speakers, view posters, mingle in the networking lounge, and even visit virtual vendor booths (we’ll have a booth there so be sure to come and chat to us!).

Topic tracks include Bioinformatics, Microbiology, Nucleic Acid Analysis and Amplification, Drug Discovery and Delivery, Lab Automation, Cell & Tissue Analysis, Biotechnology, Autism Research, Cancer Research, Immunology, Clinical & Diagnostic Applications, Chemistry, Neuroscience, Protein Expression, Genetics and Genomics. With all of these choices, you’re bound to find alot of interesting stuff, no matter what your research interests are.

Enter the lobby
Your conference experience begins when you log in and enter the lobby of the virtual convention center. This is a 3D envirornment from where you can feel the conference vibe and access all of the available options, just like in a traditional conference, although you’ll have to supply your own coffee and cookies.

Speeches, talks and posters
The lobby houses aisles containing hundreds virtual booths, each of which houses live keynote speeches or talks/posters that are going on continuously for the three day event. Enter any of the booths at the advertised time to watch a live talk, then hang around afterwards for moderated text chat with the speaker. Immediately after all talks and presentations are given live, they’re then available on-demand (for free) to come back and see later, so no need to worry if you miss something.

Network
Networking is a vital aspect of any conference, so Bioconference Live has a Networking Lounge where users can congregate to meet with their peers or network with other professionals in their field, then use the Live Chat feature to discuss industry issues and news, or just have a conversation.

Visit the exhibit hall
You can then visit the Exhibit Hall and explore new technologies.  Companies are divided by specialty or floors, so you are able to see all the vendors within your area(s) of interest in one central location.  Each vendor has their own “virtual booth”, where you can view their commercials or video uploads, peruse their pdf’s (adding them to your “briefcase” if it is something that you would like to refer back to later or download to your PC), and even Live Chat with a representative from their company if you have any questions or would like to request additional information. Suzanne and myself will be at the Bitesize Bio booth and we’d love to see you there.

Register now, win a prize
You can register for Bioconference Live by clicking here. Once registered, you can enter a prize draw (with prizes including a Macbook) by telling three friends about the conference. I’m really looking forward to this event, and hope to meet you there.

http://bioconferencelive.com/

Not only have we lasted, but we’ve thrived. Addgene recently stored its 10,000th plasmid, and our hard-working technicians are sending out nearly 200 plasmids a day to labs all over the world.

Not too shabby, but to maintain its position as the place to go for plasmid archiving and distribution, Addgene has added some new features. One is very simple and is aimed scientists searching and requesting plasmids: the flame.

After having to answer countless e-mails from researchers who were concerned about ordering a plasmid that was not field-tested extensively, we decided to just let people know which plasmids were the most popular.

A plasmid with a yellow flame has been ordered at least 20 times. After 50 requests, the flame turns red; after 100 requests, the flame turns blue. In case you’re curious, our most popular plasmid is Didier Trono’s pMD2.G (a lentiviral packaging vector), which has been requested over 1,200 times. That’s what I call field tested. Click here to check it out.

Don’t know where to start? Don’t worry. Here’s my advice.

First things first. Be sure you remember what you have already communicated about your salary; consistency is key. If you are working with a recruiter, be sure they know what salary expectations you have been communicating and be sure you know what the recruiter has been relaying to the company.

Second, it’s important to be paid fairly, but it is more important to get a job you are happy with; if you think it’s fair, it’s fair. I strongly caution against web sites that purport to provide “accurate” salary information. Each person and each position is unique.  The skill sets possessed and required are different and a “fair” salary will vary dramatically.

Third, before you can negotiate with a company, you have to know what your expectations are - we find that many people don’t have a good grasp of their own expectations and it can be disastrous during this critical phase.

I recommend knowing your “Absolute Bottom” - the number which you would say “no” to if it was literally one dollar lower. This is a very important number to know. The longer you are looking for a position, the lower this number may be, and that’s okay.

This is about the position being fair to you. From here, I would find the “I think that’s fair” number and lastly the “holy cow!” number. Armed with these three numbers, you can confidently negotiate salary and know how hard to push.

If the company offers you a number below your “Absolute Bottom” - tell them that you were expecting something in the “I think it’s fair” to “holy cow” range. You have nothing to lose because you are not going to take the position at this compensation level.

If the company offers an “I think it’s fair” salary, you need to decide if it’s worth pushing back and potentially losing the offer. A great way to approach this is simply to ask “Is there any wiggle room in this offer?”. If the answer is no, thank them and tell them either you will accept it or you’ll get back to them within 24 hours (if you know you are going to accept, there is nothing wrong with telling them at his point).

If the company offers a “holy cow” salary, tell them thank you and you cannot wait to start!

Finally, if you are anything less than thrilled with the offer, I would encourage you to sleep on your decision.  Give yourself some time to consider it and don’t act impulsively.  The offer will still be on the table tomorrow and you will feel more confident after having carefully weighed your options.

She laughed but that idea stuck with me. Current treatments are banking on the idea that the cancer is going to replicate faster than the normal tissue and will therefore be more sensitive to drugs that target proliferative processes. An example of such drug is taxol, a common and potent chemotherapeutic agent that blocks progression of mitosis thus inducing cell death.

For years, I have thought that the ideal treatment for any disease is to exploit what separates that disease from the normal tissue. It is the reason that cancer biology as a discipline has exploded in recent decades but has yielded very few distinguished targets. The personalized cancer therapy we have been promised is yet to be delivered. A new family of drugs is bringing us closer to that goal by building on years of basic biology research and targeting one of the most abundant nuclear proteins, PARP-1.

We have known for a long time that PARP-1 is important in a variety of cellular processes from replication to transcription and we are still in the process of identifying various PARP family members.

One of the most common activities of PARP-1 protein is its role in the base excision repair process that repairs single-strand breaks that occur on the DNA as a product of normal cell metabolism. If unrepaired, single-strand breaks can stall replication forks and create single-ended double-strand breaks, a highly toxic lesion repairable only by homologous recombination. Homologous recombination is one of the most commonly inactivated repair mechanisms in cancer, especially in breast and ovarian (BRCA1 and BRCA2 mutations) as well as prostate tumors. Inability of a cell to repair a single-ended double-strand break will lead to activation of apoptosis and cell death.

The pathway of oxidative damage to single-strand break to double-strand break to homologous recombination is now being used to selectively kill cancer cells. A cancer deficient in homologous recombination is going to be more sensitive to PARP-1 inhibition and formation of single-strand breaks than normal tissues.

In normal tissues, inactivating PARP-1 will produce single-strand breaks that will get converted to double-strand breaks during replication but with active homologous recombination this damage will be easily repaired and the cell will survive and continue proliferating. In BRCA2 mutant tumors, for example, inactivating PARP-1 will also produce the same type of damage but at the last stage the cell will be unable to repair it because it lacks the homologous recombination activity. Thus, inactivating PARP-1 will selectively kill cancer cells but not normal tissue, providing a silver bullet for that type of tumor.

A recent report in New England Journal of Medicine described this exact effect in BRCA2 deficient tumors during a phase I study of AZD2281, a highly selective and potent PARP-1 inhibitor.

As a scientist, I hope that soon we can hear of more therapeutic success stories that came from basic research foundations. Judging by published data, we have certainly been productive and it is now time to assess which of those avenues will be crucial as we move forward to develop new treatments that will be more potent and yet have fewer side-effects than what we currently offer.

It would be wonderful to have a treatment that would be like a skilled carpenter: able to go into a house, take out the broken window and leave without anybody waking up.

Reference:
Fong, et. al., 2009. Inhibition of Poly(ADP-Ribose) Polymerase in Tumors from BRCA Mutation Carriers. NEJM; 361: 123-34.

And when you spend that much time with people, bench to bench, for anywhere from 3-6 years, you really need to learn how to get along.

Usually, after some time, most people learn the rules of lab life and are anxious to become part of the group. When people don’t get along, there are usually specific reasons or actions of the offender that is causing the boat to rock. What are those things that can make everyone want to throw you off the ship? I came up with 10 ways people piss off their co-workers in the lab.

Feel free to add on some of your pet peeves of lab behavior.

1. Using up all of a common lab stock or buffer and not replacing it or re-ordering it.
2. Using someone else’s reagents without asking. If you contaminate them, then you are in double trouble.
3. Not doing the dishes, or whatever lab chore you are assigned, when it is your turn (or doing a poor job of it).
4. Talking too much- especially when someone is setting up PCR or other reactions that requires focus. That person is wearing ear buds and an ipod for a reason!
5. Contaminating the lab. Repeatedly. Making a mistake once is one thing but if the lab is constantly needing to be wiped down of radioactivity or other hazardous chemicals, people will start to wonder if you are cut out for the lab.
6. Hogging a community computer- especially if you are not using it for work!
7. Coming in late and leaving early: Every day? This is surely only acceptable if you are writing your thesis.
8. Abuse of shared equipment- if someone screws up the only high speed centrifuge on the floor, no one can get anything done.
9. Not signing out shared equipment and then using it when it was signed out by someone else.
10. Not giving credit when it is due- or taking credit for someone else’s idea. In a lab meeting, if someone gave you a great experiment to try and it works, let everyone know. If you don’t, don’t expect anymore help from anyone else.

This is a short list of the ten peeves that came to mind first. What does your co-worker do that you wish they would not? What is your #1 lab rule?