Archive for the ‘DIY’ Category

A quick and dirty microbiology attempt

July 3, 2012

I recently felt compelled to do some microbiology work.  My goal was to make a crude growth media for microorganisms, swab my mouth for these buggers, and spell my name across a few petri dishes.

edit:  Forgive the bright spot from my lamp, it was the only way to get the letters to show.

J

Above you can see the letter “J”.  If you look to the right and bottom of the J you can see clumpy looking gunk – that is egg chunks.

I started out by boiling 300 g of sliced potatoes and 1 raw egg  in ~500 mL distilled water for 30 minutes.  Next I filtered the solution through numerous coffee filters and then filtered the solution a second time through a 0.45 micron membrane.  10 g of sucrose and 10 g of agar-agar (hardening agent) were added and stirred for ~10 minutes.  Next I sterilized the media and some glass petri dishes (~$1.50 per dish) in a pressure cooker for 25 minutes at 15 psi.  After sterilization I poured the media into the dishes and allowed them to cool and harden.

My open dishes

My open dishes

After I finished sterilizing the media I noticed the solution had clumpy-chunky junk floating all about.  I assume this was leftover egg that got through the filters, though it could also have been impurities from the agar-agar (least likely of the two).

I used a cotton swab to collect bacteria from my teeth and gums and I spelled my name (a letter per dish) across the plates using the swab.  On one plate I put nothing and on the last plate I spit into it to cover the whole thing.

e

e

I was happy to see that everything worked perfectly.  The dish with no bacterial additions had no growth.  The dish I spit into had growth all over and the dishes I wrote letters on only grew in the shape of the letter itself!0

s

s

Unfortunately condensation leaked all over the plates when I flipped them and blocked some letters from being photographed.

For future experiments I bought a “fitness multivitamin” that contains all of the amino acids and I am going to try and use it as an egg/tryptone/peptone replacement.

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Lab grade vs. home grade electrophoresis buffers

June 8, 2012

(note:  wordpress sucks so it screwed up my images some.)

I wanted to find a cheap way to make electrophoresis buffer from easily accessible ingredients and there were two pieces of knowledge that led me to this experiment.  First I had read about some labs using sodium borate (SB) buffer because it gave great results.  Secondly, I knew that borate and boric acid were easy to find at stores in the forms of roach poison (boric acid) and borax (sodium tetraborate).

Originally I had intended to compare a large number of buffers; TAE, TBE, SB (molecular biology grade), and SB (home grade).  Due to my own personal shortages of lab-grade materials I had reduce down to a comparison between TBE, SB (home grade), and SB with EDTA (home grade).

All gels were 1% agarose.  Gel images were made with Foto/Phoresis I transilluminator and gel images were recorded with an iPhone 3GS camera.

Tris-Borate-EDTA (TBE)

Purchased from the “Online Science Mall” as a 5X concentrate.  Without a doubt this worked better than the buffers I cobbled together.

 

SB Buffer (home-grade)

SB buffer made by adding 1.91g of Sodium borate decahydrate (Borax by 20-mule team) to ~800mL distilled H2O (Target), pH was adjusted with 0.4M Boric Acid (Roach Away by Enoz) and then diluted to final volume with more distilled water.

SB Buffer with EDTA (home-grade)

Image

A 600 mL aliquot of the SB buffer, made previously, had 1.2 mL of 0.5 M EDTA (molecular grade) added to reach 2mM final concentration.

I decided to test the SB buffer with EDTA to see if there were any nuclease related problems occurring.  2/3rds of the way through the project it occurred to me I was using Target brand distilled water instead of nuclease free water when mixing the 2-log DNA.  It just simply slipped my mind.  So this third gel buffer test was done with EDTA and nuclease free water to see if the results would be drastically different or not and they were not.  I believe any differences in the images is due to my lack of a proper gel documenting system.

Given the choice between the three buffers, TBE is the superior.  I do not mean to say that one could not use the SB home grade buffers and get usable results, but rather they just are not as good.  Being that store bought TBE and TAE are not crazy-expensive and buffers can be reused a couple of times, there is not a lot of impetus to use home-grade SB buffers.

My results for the SB buffer tests do not even come close to mirroring the results from the Brody paper I linked earlier, I suspect that my preparation or formulation of SB may be flawed.

While unfortunate that I couldn’t test more buffers, I was happy to figure out that TBE will likely work for my PCR projects in the future and if I am ever mismanage my supply of TBE I now know I can make up some SB buffer to get by.

Build a pipette stand from PVC pipe for $3.47+tax

March 18, 2012

Cross-posted here and at Citizen Science Quarterly.

I wanted to purchase a pipette stand but it seemed like such a frivolous purchase. Why should I spend between $20 (eBay) and $60 (venders) to hold my pipettes vertically? As much as I wanted a pretty acrylic stand I just could not pull the trigger and buy one (I’d rather buy DNA ladders). This post will briefly go over how I built a pipette stand out of PVC pipe.

These instructions are for a linear pipette stand. The circular/revolving stands are an annoyance and an example of bad design. If you are suffering through one of the revolving stands I urge you to abandon it for a linear stand.

Tools

Saw or PVC cutter – Either work but the PVC cutter is easier to use and costs $10-20.

Marker and Tape measure

Materials

(above) All of the supplies needed for the stand.

PVC Pipe

Pipe is usually sold in 10 foot lengths. Most stores will cut it down smaller for free if asked. Some stores even sell shorter segments. Cut the pipe into four 8″ lengths, two 6″ lengths, and six 2″ lengths.

PVC Fittings

Purchase four T-shaped fittings and six right-angle fittings.

Cost breakdown

T-shaped fittings (4) = $0.80

90deg fittings (6) = $1.02

½” pipe (10 feet) = $1.65

Total = $3.47

Instructions

The construction of the stand is simple. All that has to be done is to cut the pipe and then assemble the structure.

As mentioned in the methods, cut the pipe into four 8″ lengths, two 6″ lengths, and six 2″ lengths.

Next connect the fittings together with all of the 2″ lengths of pipe as shown below.

Now add the 8″ and 6″ lengths of pipe – the stand is now complete.

(above) A view of the stand from the bottom.

(above) Fully assembled stand.

(above) A view of my home-lab with the finished pipette stand.

Ceiling fan centrifuge.

March 11, 2012

I needed to pellet a saline solution containing cheek cells for genomic DNA isolation but I did not have a centrifuge that could hold 50mL tubes.  The solution was to tie the centrifuge tube to my ceiling fan with shoestring.  The ceiling fan centrifuge was ghetto, scary-looking, and effective (it had 3 different speed settings!).  I made a video, check it out below.

Before using the fan I first tried to spin the shoelace (with tube) by hand – but this did not work out at all as the solution kept getting stirred.

 

 

 

 

DIY Tube Holder for Vortex Mixers

January 20, 2012


The protocol I use for collecting human DNA samples requires tubes to be vortexed for 10 minutes. Standing around for a sixth of an hour is not my idea of fun so I decided to get a foam tube holder. Unsurprisingly a piece of foam with holes in it costs 50 dollars. As per my usual I wanted to be a cheapass and thus I built my own foam tube holder with some things I had lying around. If you want to see the DIY tube holder in action, watch the youtube video below and scroll further down for instructions for building your own.

Guide

Compatibility

Before attempting this guide make sure your vortexer will work with this method. Check the type of head piece present on your vortexer, consult the diagram below for an example of two different types (#12+13 and #14). This guide works for vortexers with head pieces that match #14. New vortexers usually come with both of these pieces and used vortexers (like the one I bought) may only come with one type.


Materials

The tools and consumables I list are not absolute. Use your noggin and substitute if necessary.

  • Tools
    • Dremel (a drill can be used for some steps but not for carving foam)
      • Sanding bits are needed for foam carving (see image below)
      • Cutting tool
      • Drill bit
    • Ruler or measuring tape
    • Hand saw
    • Marker
    • Compass (optional)
  • Consumables
    • Foam block
    • plastic box (I used a large pipette tip box that was 4″x5″)
    • Rubber bands


These are the two dremel tips I used to carve the foam.

Instructions

Two types of modifications need to be done to the plastic box. First, a hole needs to be made in the center that will allow the vortexer head to poke through. This alteration will prevent the tube holder from wobbling off of the vortexer head. For my vortexer the size of the required hole was 1-1/4″. The second modification is the addition of 4 grooves to the top of the box so that the rubber bands do not slip.


Prior to drilling and cutting grooves. Yellow circles indicate approximate target locations for the grooves.


After drilling and cutting grooves


Close-up of the grooves


How the box fits onto the vortexer head.

Now it is time to carve the block of foam. My box was not perfectly square; the top of the box (the opening) is larger than the bottom of the box. This odd shape can be ignored but do make sure to use the top of the box for determining the foam block size. I made a block that was 4-1/4″ x 5-1/4″ x 2″. It is very important that the width and length match or slightly exceed the size of the box so that the foam sits tightly inside the box.


My block


My original block was too tall. Rather than cut grooves in the foam for the rubber bands and have a tall block (which is preferable for large tubes), I decided to cut the foam down so that it was flush with the box.

Carving and cutting the holes in the foam can be frustrating. Make as few cuts and holes as possible because with each tear the foam becomes more likely to get caught on the Dremel bit and then it will twist and tear the foam block. Also note that the edges of the foam block are likely to get caught by the Dremel bits.

I used sanding bits to make the holes you see below. I started with the cylindrical dremel bit I pictured in the materials section. I went into the block about 0.5″ with the cylindrical drill bit (this made the cleanest looking hole opening) and then I switched to the conical bit which I used to go straight down to the bottom of the tray. Use tubes to test each hole you make to ensure they fit. A snug fitting tube is better than a loose fitting tube.


When the Dremel grabs the foam and twists some areas of the block will be torn.


Another view of the finished block.

All that is left is to attach the box to the vortexer and for that we just need rubber bands. The way the rubber bands rest on the vortexer head and on the box is important, consult the images below.


The first rubber band is hooked under the left side of the vortexer head and hooked over the right side of the box.


The second rubber band is a mirror of the first rubber band.

That is it, the attachment is finished.


Before


After


Glamour shot


If you build a vortexer attachment send me a picture and let me know how it turns out.

Quick Update:  The more practice I had with the foam, the better I got making a cleaner/nicer looking piece.

How to build a $21 gel box.

October 22, 2011
My box

My box

My results

Citizen Science Quarterly asked me if I would like to blog at their website and I said sure.  The first thing I blogged about was how to build a gel box for 21 bucks.

Click here to read about it.

LEGO electrophoresis box, as finished as it is going to get.

October 2, 2011

I kinda-sort finally finished the LEGO electrophoresis box.  Saying  ‘finished’ is a half-truth of sorts because I decided to abandon the nearly functional gel box.  Everything worked out but a few issues made the LEGO gel box more effort than it was worth. Keep reading for the details.

Final box

In the beginning….

When I originally set my mind to making the LEGO gel box I decided to test whether it was true that acetone would melt the ABS plastic (which the LEGO’s are made of) and thus fuse the bricks together and I wanted to test how nice of a gel could be cast.  The image below shows a still molten agarose gel being cast inside the rotatable gel mold.  To my delight, it was true that acetone would meld LEGO’s together and the LEGO’s worked great for casting a gel.

The prototype

I decided to class things up a bit and make the second version of the gel box all clean and spiffy looking (see image below).  The main impetus for using a single color LEGO brick was to avoid the color smearing that was inevitable.  The application of acetone melts and leads to smearing of the different colors.  In short, it looks dirty.

Black is the new black

Some assembly required….

The gel box can be assembled in an infinite number of ways and each person can go about things however they want.  The box I built was 12 LEGO pips by 28 pips and was 4 bricks tall.  Being the second attempt at a LEGO box I decided to step things up a noth and add 4 small protrusions on the inside of the box to hold the gel mold in place.  These protrusions stick out one LEGO pip and they do not obfuscate the electric field as the gel mold already blocks this area.

I entertained quite a few electrode placement variations before ultimately coming up with the design you see above.  I originality wanted the cables to plug into the side of the box, but the size of the banana plugs and the stubbornness of LEGO bricks to be drilled neatly prevented a side mount.  The LEGO plates I ended up using rest on top of the gel box and hang over it.  Take a look at the image below.

No electrodes

The image above is the gel box without electrode flaps and the image below is the box with electrode flapss.

With electrodes

Creating the electrode holders was easy.  Start with a LEGO plate that spans across the gel box and decide where to place the hole.  Once the spot has been chosen, drill a hole by starting with a small drill bit and moving up sizes until the hole is the needed size.  Starting out at the largest bore size will likely cause the plastic to crack or warp.

My reaction to drilling LEGO’s surprised me. -I was filled with a combination of childhood nostalgia and irrational love for LEGO which urged me to quit!   I nevertheless worked past my irrational love and generated the part you see below.

Drilled hole.

The next step is to attach the bannana jack.  The jack I bought attached to the LEGO plate by a screw on the underside (no gluing required).

Banana jack.

Electrodes attach to the jack similar to how the jack attached to the LEGO plate – via a screw pinching the material.  To make the electrode I stripped some wire and twisted it to form a cable.

Electrode

Once the electrodes are set-up its time to work on some finishing touches.  First, a gel tray or cast is needed.  The image below is the prototype tray I made, I chose this image over the new one I made because it is easier to see this one (on account of my camera being unable to image the black colors).  The mold is  a few LEGO plates, flanked on two sides by 1 pip wide bricks and covered with smooth plates.  The molds I built were 8 pips by 8 pips.

Mold

Building a lid for the gel box is critical, that is unless being electrocuted sounds appealing (note, the electricity could kill someone).  I lacked any single LEGO plate that could cover the entire span so I made my lid two LEGO plates thick and staggered multiply pieces so I could span the gap.  I lined the top of the gel box with smooth-topped plates and a couple normal plates so that the lid would attach but be easily removable.

Final box

After the box is assembled it is time to  weld all of the bricks.  When I chemically treated the gel box I used pure acetone and wiped all sides of the gel box.  I avoided welding the lid because there was no need to make it waterproof.

As an aside I recently read an article from MAKE magazine that gave a suggestion for making ABS glue.  They suggested adding ABS plastic flakes or pieces to a small bottle of acetone and letting the plastic dissolve over night.  The addition of plastics supposedly helps with the combining of the ABS plastic later on.  I have never tried this, but I will if I ever need to chemically weld plastics.

After acetone was dried up I performed a water test.  To my delight, the box held in water almost perfectly.  There were only three leaks present and all of them were at the junction between the walls of the box and the yellow base-plate.  I decided to be a cheapass and not purchase silicone to seal the bottom edge and I now regret this.  Instead of the silicone, I used a thick bead of PVC glue all around the bottom.  This was a bad idea.  The PVC blocked up two of the holes just fine, but the third was not stopped.  The problem was not the PVC glue sucked at sealing the bottom but that I could not cut out the PVC glue and re-seal the problematic spots (PVC glue melds with plastic).  When I tested the box after PVC glue treatment, the remaining leak was worse than the three original leaks, combined.  The problem was damage from transportation.  Going from my apartment to the lab led to the base-plate becoming even more loose and made the leaking worse.

The leak was the straw the broke the camels back and made me abandon the project. In my opinion,  if moving the gel box around could easily lead to leaks forming then the box was not worth the effort and I should probably just use a Rubber-Maid container or something.

The last problem I had was with chemistry.

The image below shows a electrochemical reaction that, while kind of interesting, was frustrating to encounter.  The twisted wire worked just fine for the anode, but the cathode  reacted to the current and chemicals.  I have no clue what exact reaction is occurring but the end result was the wire becoming a striped red and green color (which I think means the wire is made up of separate smaller wires wound together, probably copper and something else).

Frustrating reactions

(-)

All done.

The following two images are of the (mostly) finished gel box.  Despite the issues I had the box worked and so I am calling this a success even though I abandoned the project at the finish line.

In operation

 

Hooked up and with a lid

While I may have given up on using LEGO’s I have no given up on building a cheap box (<$10 a box).  Below is a preview of the new gel box I am working on and I will blog about it later.

Cheapo box

 

 

 

Update: Microscope incubator

September 21, 2011

After putting the DIY microscope incubator I recently built through its paces it has come time to revise and improve (evolve?) it.  Aside from enlarging the chamber just a tad I have figured out how to more elegantly integrate the hair-dryer into the incubator.  There should be a significant reduction in waste heat now.

Ahhh yea. (Top view of the incubator)

Yea…I am going to need a dremel.

September 20, 2011

Ugh, wow that is bad.

I tried to carve a gel comb out of a piece of LEGO.  Scissors, pliers, and a razor yielded a result that is less than satisfactory (see above).  I was really hoping to avoid buying a Dremel but the outcome of my first gel comb has convinced me to purchase one.

LEGO electrophoresis box update.

September 19, 2011

I spent some time this weekend on improving the LEGO electrophoresis box.  Below are two images.  I should be done with the whole thing and testing it some time this week.

One thing holding me back is that I have been having a lot of problems soldering cables.  I have never soldered before and I think the iron I bought is defective.  Eventually I will work my way through the problem.

Box with cover.

Box without cover