Electrophoresis Kits

[Pm706Narrogin]

My confession is I tend to wear my lab coat and gloves even when doing innocuous things not cause I'm super safety conscious but just because I'm a klutz who's liable to do this exact thing at any moment

[bigmack]

Google AI said I could freeze my dyes to stop them going mouldy . It said they wouldn’t freeze due to the glynerin

Hmmmm

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[Pm706Narrogin]

Ah I see the machines can't replace us just yet

[bigmack]

Had a play yesterday .
This is a mixed bag . Made with Agarose and using TBE buffer .

The left hand trays are using PM706 Muscular dystrophy scenario . The big pink splodge happened when I loaded the dye . Not sure if I stuck the pipette through the well bottom or if the bottom tore out when I removed the casting comb .

The right hand single tray was an experiment . My wive says she loads the trays ( gets the students to do it ) before pouring in the electrolyte . So I wanted to see if it would work .
Also these are the four dyes I have made from scratch .

The bottom middle are the same four homemade dyes but loaded with the electrolyte poured in to compare with dry loading .

The top middle is the Southern bio dyes .

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This is how it finished .
The Muscular dystrophy scenario at the bottom left was amazing. The one above not so clear . I suspect all the residual dye that floats around got pulled up from the bottom tray and sits under the top tray .

Seems the home made dyes work very well . There didn’t appear to be much difference between dry loading and wet loading the trays .

The Southern bio dyes worked well as to be expected but as usual the green dye moved backwards and off the end of the tray .

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I tried it again , but this time using just plain Chem supply Agar ( not agarose ) and 0.1% Sodium Hydrogen Carbonate buffer and electrolyte .

This time I added the xylene cyanol from the SB kit in my lineup and you can clearly see the different weights of the five dyes and how they move differently .

I also repeated the Muscular dystrophy scenario to see how it compared to doing it with the expensive Agarose and TBE buffer .

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My take home from all this is that there is a slight improvement using the Agarose and TBE but it is not massive .

I also don’t like the stacked trays . There seems to be too much smearing on the top tray . Sure , it doesn’t interfere with the results , it just looks yuk.

I am pleased I have decided to get and use my own dyes . I must say I was hesitant at first . So thanks to PM706 for sharing all their knowledge .

Also , Xylene cyanol was not actually needed in the scenerio I used . Incidentally , I have some coming but could have probably not needed it . The only place I could find it in Australia was Merck . Its cost $200 including freight. But we should be right for many years now .

Also to note .The reason I tried dry loading the wells is because if you have 6 groups , each can take a tray and load the dyes at their work station simultaneously . Then bring them back and place them in the tank and pour in the electrolyte .

Other wise , it could take a while period for each group to do it one after the other . And by that time , the first few trays start to do funky things . I guess this is the beauty of having multiple single tanks , but I have what I have .

[bigmack]

Ah I see the machines can't replace us just yet

Not for a while yet .

Google AI also just told me that Orange G will travel fastest and furtherest because it has a lower molecular weight than Tartrazine .

It also said that xylene cyanol was the exception to the rule and it will travel faster than Rose bengal because of its molecular shape .

Well seeing is believing .

All this is clearly wrong as Tartrazine was way ahead of Orange G and xylene cyanol was the slowest of them all .

Apparently , it is the overall charge of the dye molecule which determines its speed and distance , and while it is sort of related to molecular weight , that alone is not the reason .

1st Yellow —— Tartazine —————————534.36 g/mol
2nd. Orange —— Orange G———————— 452.38 g/mol
3rd. Purple ——-Bromophenol Blue ———670 g/mol
4th. Crimson——Rose Bengal———————973.67 g/mol
5th. Blue ————Xylene cyanol——————538.61 g/mol

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[bigmack]

I setup the Pm706 Tuna /whale meat dyes today and had a try . Used the Xylene Cyanol from the SB dyes kit . ( mines still coming from Merck)

Again , using plain Agar and bicarb buffer .
When it comes to dyes , there appears to be no reason to use TBE buffer at all . It is needed purely for real DNA samples .( which we aren’t using ) …… and it’s expensive

I use 1g of Sodium Hydrogen Carbonate in a Litre of distilled water . Use it to make the Agar and pour in the tank as the electrolyte .
1% W/W Agar . ( this is from Southern biological Carolina Dye instructions )

I also discovered if I put the trays in the fridge for an hour after casting , the cooler gel makes the combs pull out cleanly . At room temp , the combs don’t come out nice and can tear out the bottom of the wells .

My kits came with 40uL pipettes . It is boarder-line too much and if the gels are not poured thick , the wells are not deep enough to hold 40uL and it tends to “pull” out the top of the well creating a smeared looking result .
My last school used 10uL . I split the difference and went fir 25uL and that seems like a happy spot .

Sorry for the waffle but hopefully , all info is helpful .
Anyway here is the results of Pm706’s “Naughty fisherman”

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My Bio teacher is really excited .

[Pm706Narrogin]

Those gels are looking great. Glad to hear the dye mixes are working out.

And yes I've only ever used the 1% bicarbonate buffer and never had any issues.

The other thing I love about the different scenarios is you often have a few kids who choose both Bio and Human Bio so both courses can do electrophoresis and even if they've done it with one class they can get a different scenario and still get something out of it. The Southern Bio kit seems like the sort of thing you see once and that's it

[bigmack]

The teacher ran it with the kids . She asked me to come and assist with showing them the process . They all git given a practice gel in a petrie dish with water to load . Then each group came and loaded their tray . Unfortunately , it took too long and the bell rang fir lunch 10min into the run so they didn’t stick around to see the results .

The teacher did, and took pics . Today they studied the pics and were able to work out the problems in the scenarios .
She was very happy .

Today , one of the students saw me in the playground and said “ Sir , that electrophoresis lesson was one of the best, I have really enjoyed and leaned from it “

[Pm706Narrogin]

Aww those are the reasons you push through all the frustrations and keep coming back. Knowing that something you do might help even one kid understand something more or ignite a spark that makes them want to choose science as a career.

But also what a glowing review for ChemTalk. You have a lab tech in WA who has since retired (full credit to Lesley W) who made these different scenarios. Then through a discussion on here that work gets shared, gets workshopped by a labbie in Queensland and enjoyed by students on the other side of the continent. How good.

[RosalieL]

Aww those are the reasons you push through all the frustrations and keep coming back. Knowing that something you do might help even one kid understand something more or ignite a spark that makes them want to choose science as a career.

But also what a glowing review for ChemTalk. You have a lab tech in WA who has since retired (full credit to Lesley W) who made these different scenarios. Then through a discussion on here that work gets shared, gets workshopped by a labbie in Queensland and enjoyed by students on the other side of the continent. How good.

I second this!!! It is such a good resource. We are hoping to do a forensics day with year 10 next term so I'm planning to use this information as well. If you have contact details for your retired lab tech, feel free to thank them from the East Coast! (I am near Newcastle, NSW)

[RosalieL]

Aww those are the reasons you push through all the frustrations and keep coming back. Knowing that something you do might help even one kid understand something more or ignite a spark that makes them want to choose science as a career.

But also what a glowing review for ChemTalk. You have a lab tech in WA who has since retired (full credit to Lesley W) who made these different scenarios. Then through a discussion on here that work gets shared, gets workshopped by a labbie in Queensland and enjoyed by students on the other side of the continent. How good.

Is there any chance you could email the files/notes you have? I tried to copy and paste but it wouldn't work...

[Pm706Narrogin]

I'll save copies from my work computer and shoot you a PM later, Rosalie.

And if anyone else wants copies of the notes let me know. I tried attaching the notes originally when I was writing it up but they're too large to be attachments on ChemTalk sadly

[RosalieL]

I'll save copies from my work computer and shoot you a PM later, Rosalie.

And if anyone else wants copies of the notes let me know. I tried attaching the notes originally when I was writing it up but they're too large to be attachments on ChemTalk sadly

I've sent you my email address via PM - Thank you!