The Art of Corner Cutting

The industrious individual is a golden tool: rare, highly valued, and gleaming with potential.

The average graduate student is a 200 pound gorilla with a micropipette duct-taped to its hand: expedient, irritable, and lacking fine motor control.

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Confident, bold, and daring. Like it or not, this is the ideal graduate student.

That said, we should not forget that even the most brutish lab wildlife gleams with the potential of an experiment well done and a question elegantly answered.

So where is the line between lab gorilla and Gautama buddha: prince of pipettes? To put it simply, the difference between the two lies in their adeptness in the art of corner cutting.

Obviously, every individual has the capability to do careful, thoughtful work and expedient, shit-tier work. As humans we often like to assign moral values such as good and bad to the quicksilver and quaint respectively, but let us for a moment consider that the entire spectrum of output quality has its corresponding spectrum of appropriate niches.

What wily and clever lab rats know, consciously or unconsciously, is that to maximize efficiency is to employ the correct quality depending on the demands of the situation.

Why did I spend an hour on Friday night making this?

Serial Dilutions of Skill

Now when we talk about quality, and in particular the quality of one’s work, we must first define standards to compare to. Taking that into account, I’ve gone ahead and split the quality spectrum into tiers and given examples below of the kind of work and worker you can expect at each.

“I’m not actually a shitty scientist I was just pretending haha” Tier

At rock bottom we find the “laser eyeball colorimetric readout” your mentor hastily taught you when he was too lazy to walk to the plate spectrophotometer. This type of work is all corners and cuts, no actual product.

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“Yup that look’s like 2mg/mL to me.” – That one grad student who leaves at 3pm every day
HEY GUYSLOOK HOW RETARDEDIAM HUR HURRRRRRRR fuck offretard JOKES ON THEMIWAS ONLY PRETENDING face white cartoon text facial expression black and white line art nose human behavior head emotion drawing font line diagram

Sadly, this is usually a grad student who knows better but has completely lost any passion for science and discipline.

Leaves samples in the thermocycler for weeks-on-end Tier

Increasing in quality a bit we find the average undergraduate’s typical output. This consists of assays where directions were technically followed but without any real forethought or care. Examples include the no longer supercoiled DNA preps that have been left on the bench for 3 weeks and an unfathomable number of unlabeled tubes that may or may not be important.

This tier also encompasses the work of those who lack motor, technical, and intellectual skill. That guy who always contaminates the LB after taking 20 mL from the 1 L bottle comes to mind but he’s far from the most dastardly criminal who resides in this cursed domain. Case in point: RNAse guy.

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Ancient representations of RNAse guy. Note the elongated head that allows for more surface area to shed RNAse from. Disgusting.

RNAse guy is the extrovert that, no matter when or where you start your RNA prep, will come up and start talking and breathing in your personal space, spewing all his disgusting RNAses into your precious sample. I’m non-ironically mad writing this because it happened yesterday. Fuck RNAse guy.

Side note: leaving your DNA at room temp really will cause loss of supercoiling and downstream losses in efficiency when used for transfections or in vitro assays.

Avoids talking to the PI / Time Wizard Tier

Another increase in quality brings us to what may be considered passable work, sometimes.

This is the dude who cruises Instagram in between flow cytometry samples but won’t take the time to use the autosampler. Experiments and corner cutting at this level resemble a gambling addiction more than science and while failure is common, occasionally this high risk-high reward strategy pays off big.

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“Gloves are the only PPE I need!”

This tier is unique in that it has a very recognizable aesthetic. Contrary to the norm, lab coats and aseptic technique are typically not employed at this level due to the student’s own perceived expertise and “secret breath holding technique” (Side note the breath holding totally does work though).

This student has been around for awhile, at least 1 lab generation and could be considered a journeyman corner cutter. When not catastrophic, the shortcuts this student takes do not have an immediately noticeable effect on the work produced. The student, however, will constantly complain that assays are, “inconsistent” and that, “science is a joke”.

To be fair, this student is right to complain that science will always be contaminated with unknown variables due to the unfathomable complexity of the universe, but still foolish because he discounts his own role in increasing the unknowns in every experiment through expedient action and insufficient planning.

Overall the most fun strategy. Highly recommended.

Lab Tech Tier

There’s not much to say about this tier. These people are actually paid to do their research and they generally complete all necessary work in an organized and orderly fashion. Sure they leave at 5pm but that’s their prize for not procrastinating until the last minute to pass their cell cultures.

Corners are not cut in this sacred domain and as a result quality is high, but this consistency comes at the cost of time and adventure.

Enlightened Buddha Tier

The enlightened student is the paragon of corner-cutting optimization: quick, fluid, and with a fairly high rate of success. This student empathizes so intensely with the mistakes of his colleagues that he learns more from their errors than they do.

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This student never contaminates their sample with RNAse because they don’t ever breath, they simply allow oxygen to diffuse into their blood by coating their body in DMSO every morning. Mmm, garlicy.

The cultured cells of the Buddha student are cared for as one cares for a friend, or for themselves, because the student recognizes that all living beings have a nature tied to his own- not of the same birth or the same blood, but of the same living essence and possessing a share of the divine. This student holds funeral processions when they bleach their high passage number cultures.

This student is an artificer of the highest quality and to explain why some corners are cut and others not would be comparable to explaining mans’ place in the universe.

The Sound of Water…

Why is this article titled “The art of corner cutting” and not, “5 TIPS YOU DIDN’T KNOW FOR CORNER CUTTING FAST | UPDATED 2020 | IMPRESS YOUR ADVISOR | FALSIFY DATA ™ | ‘Free… your work… from the British Empire” -Ghandi“? Well, it’s because learning when to focus on speed and when to be meticulous is too complex an equation to compute from any set of written instructions.

Arts are arts because their performance defies the conceptual boxes of language. The explanation of how to dance can’t give me the skill of dancing any more than the statement, “The tea is hot”, can give me the feeling of hot tea on my tongue. Only dancing and drinking tea can do those things.

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Luckily, humans possess the often underestimated tool of metaphor, whereby I can transfer a conceptual image (a myth) to you that conveys the performance of an art. Additionally, I can give you examples of good and bad corner cutting so that the complex patterns that underlie successful skimping can be transferred to you.

What follows is an attempt to condense the sound of water to a, “plop!”

Types of Corner Cutting

I categorize the types of corner cutting into 2 main categories: insufficient planning and improper execution. As we will see, while the implementation of an idea can (and almost always will) diverge from the plan, the lack of a plan entirely cascades into exponentially more trouble than is worth.

Improper Execution

Reducing & Omitting Procedures

The decision to omit a step should be made based on these criteria: predicted influence on outcome (perceived importance) and cost-benefit of skipping.

When designing an experiment your brain will unconsciously assign an “importance value” to each step. Calculating such a value empirically is not feasible so this is a contributing factor to the whole art concept.

As an example, skipping the addition of IL-4 to a B cell culture is a bad idea because survival cytokines are extremely important for survival (obviously) and the time cost of the procedure is about 30 seconds. The risk is a crashed culture and 2+ weeks lost.

On the other hand, if you go to your fridge to find you only have half the amount of IL-4 necessary for the culture volume and have to decide between bleaching half the cells or taking the risk on halving the cytokine concentration, a reasonable argument can be made for skimping.

If you go down this path, err on the side of reducing rather than eliminating steps and always document your modifications. Sometimes less turns out to be more!

Saving Time or “Lust”

Now I get it, graduate students have the schedule density of world leaders without any support and sometimes time needs to be cut from somewhere. Let’s approach protocol condensation with harm reduction in mind and with the disclaimer to use your best judgement based on the downstream application. Here’s how to do it:

Do

  • Reduce incubation times by up to 50% when working with nucleic acid when a purification step follows
  • Use fewer higher volume washes over more lesser volume washes (ensure >2000-fold dilution factor)
  • Skip incubations when transforming bacteria with high quality plasmid
  • Solidify agarose gels in the refrigerator instead of at room temp
  • Dirty passage cancer cell lines (Spin down, resuspend, plate w/ out wash)
  • Estimate PCR annealing temps instead of doing a gradient (Annealing temp usually = half way between melting temps of both primers)

Dont

  • Deviate from protocol on any primary cell culture
  • Omit addition of any reaction components in in vitro assays
  • Skip ANY step or precaution when working with RNA
  • Fail to record protocol modifications
  • Freeze/Thaw protein stocks any more than necessary
  • Treat main stocks of reagents without care (you will regret it later, trust me)

Pausing a Protocol Overnight or “Sloth”

This lesser sin is one of the most tempting ways to cut corners. When a protocol is 12 hours long it’s reasonable to try and space it over 2 days to reduce fatigue.

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What really matters here is what kind of biomolecule or assay you are working with. Remember that average stability is as follows: DNA >>> Cell Culture > Protein >>> RNA

DNA is basically bomb-proof but improper storage may result in conformational change (i.e. loss of supercoiling) or degradation if nucleases are present.

In cell culture assays there isn’t usually any good pause points and you will see the ramifications of your sloth in the results. The best place to call it is just prior to any kind of nucleic acid or protein extraction from a pellet, in which case you can just flash-freeze the cell pellet and keep it in the -80C overnight.

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Purified protein is the chill guy from California who takes it easy under favorable circumstances but loses his shit when things get a bit warm. Clarified lysate and purified protein / columns can be stored at 4C overnight for most proteins with the addition of a protease inhibitor, but occasionally our polypeptidic friends will take the easy way out and degrade, or more likely, precipitate. Don’t expose to room temperature.

RNA does not want to exist in this world. Water hydrolyzes it at every temperature including -80C. Don’t skimp here or you’re in for a world of RNAse activity.

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Add lots of murine RNAse inhibitor to your stocks if you wish to store them overnight at 4C but if you’re wise you will do an ethanol preciptation and store RNA as a pellet under ethanol at -80 C. Yes it’s a pain in the ass. No it will not phase into the void under these conditions. Ever.

Insufficient Planning

Winging It or “The Free Bird”

The “Winging It” strategy is the 9th circle of Hell (Treachery) in Dante’s Inferno. You will find yourself frozen in place by gusts of chaos and uncertainty generated by what will seem to be a literal demon.

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A few months back I decided to check my activated primary B cell cultures for the presence of exosomes with no prior knowledge about exosome isolation or characterization. How is it then that I ended up sitting at the flow cytometer at 2AM alone, afraid, and exosomeless? Well, after 6 hours of culture supernatant ultracentrifugation the effects of my lack of premeditation became apparent. It turns out my instrument can’t detect particles as small as exosomes without a suite of modifications and degassed, 0.2 um filtered buffers. Oops.

My undergrad and I call these types of experiments “fuck it expeditions”. Crass? Yes. Accurate? Absolutely.

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Deciding to completely wing an experiment, going in with only a hypothesis and no plan, is analogous to trying to talk your way out a speeding ticket when the cop is deaf: you better be quick on your feet. The key in this type of experiment is to make like Lewis and Clark and map the territory effectively.

Recognize from the beginning that your chances of a reproducible success are nil and that the gold to be gathered here is in how what you learn will inform your future experiments. Focus on taking detailed notes and when problems arise, which they will, write them down so they don’t take you by surprise in the future.

Don’t spend more than a day on this type of experiment, and definitely do it on a day when your PI is not present.

In my case I learned that any exosome investigation will require a team of dedicated undergrads to do all those ultracentrifugation steps for me.

Insufficient Planning – The Partial Wing or “Mad Libs for Scientists”

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Partially winging an experiment means plunging into the depths with the mere outline of a battle plan. They say the difference between madness and genius is the outcome of the ideas and this strategy is the same.

Success here is dependent on the length of incubation times associated with the experiment. This is because as you proceed through the steps you will need to be constantly readjusting the strategy to account for the uncertainty in the unplanned steps, and this takes time.

As progress is made and experience gained the correct course of action will become obvious, but if you have insufficient time to consider your options or gather reagents you will pay the price in delays. Remember this: note carefully all the alterations in protocol you make.

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