Lab techniques page

[drteeth]

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I wanted to start a page that listed all possible lab techniques that might be necessary for the DAT. I know lab techniques such as distillation are commonly asked questions so please post as many techniques as you can think of and explain what they do and how they work.

Labs have never really been my thing, but Ill do my best to begin:

From Kaplan:
Simple Distillation: Used to separate liquids that boil below 150 degrees and at least 25 degrees apart.

Fractional Distillation: Used to separate liquids that boil less then 25 degrees apart.

Vacuum Distillation: Used to separate liquids that boil above 150 degrees and at least 25 degrees apart.

Centrifugation: Separation of particles in solution based on density. More dense particles settle to the bottom while less dense particle stay on top.

Filtration: Separates solids from liquids.

 

[predentlsu1]

Chromatography and Electrophoresis
I had a question about Chromatography on the 1st DAT I took. The basis of the question made you differentiate between ion-exchange chromo, size-exclusion chromo, and affinity chromo.
Questions about what happens to a molecule when it is placed in an electric field (migrate to anode/cathode etc.) also seem to be common.

 

[Lonely Sol]

Chromatography and Electrophoresis
I had a question about Chromatography on the 1st DAT I took. The basis of the question made you differentiate between ion-exchange chromo, size-exclusion chromo, and affinity chromo.
Questions about what happens to a molecule when it is placed in an electric field (migrate to anode/cathode etc.) also seem to be common.

I was wondering the answer for the last question you asked! Wouldnt that depend on the charge of the molecule. If it is negatively charged then it will move towards anode if negatively charged and vice versa? So, do they give you the charge on the molecule specified in the question?

 

[drteeth]

Chromatography and Electrophoresis
I had a question about Chromatography on the 1st DAT I took. The basis of the question made you differentiate between ion-exchange chromo, size-exclusion chromo, and affinity chromo.
Questions about what happens to a molecule when it is placed in an electric field (migrate to anode/cathode etc.) also seem to be common.

OK, so I know how affinity chrom works but Im not sure about the others. Could you follow the format that I was using in my above post and clarify these methods and what theyre for please...

Thanks

 

[predentlsu1]

The following 3 types of Chromotography all fall under Column Chromo

Ion Exchange Chromo: uses beads in a column, that are coated with either positive or negative charges to attract compounds with opposite charge. Ex. beads with a (+) charge will bind compounds with a (-) charge while (+) charged molecules fall through.

Size-Exclusion Chromo: also uses beads, however these beads have tiny pores. Small molecules enter these beads and their movement down the column is slowed. Large molecules move around the beads and thus travel down the column faster.

Affinity Chromo: these columns can be customized to bind a substance of interest. If you desire to pull a certain substance from a solution, you would coats the beads of the column with something that binds that substance tightly. This substance would be bound within your column while the rest of the solution is washed out.

Heres a couple of links for detailed explanations about chromtagraphy
http://en.wikipedia.org/wiki/Chromatography
http://www.rpi.edu/dept/chem-eng/BiotechEnviron/CHROMO/chromintro.html

And yes I was refering to how molecules move in an electric field based on their charge.
ANions are attracted to the ANode
CATions are attracted to the CAThode
There are 3 Types of Electrophoresis:
1) Agarose Gel Electrophoresis: used to separate pieces of DNA based on size
2) SDS-Polyacrylamide Gel Electrophoresis: Separates proteins on the basis of mass and not charge.
3) Isoelectric Focusing: Separates proteins based on charge
At low pH [H+] is high, therefore at a pH<pI proteins will be (+) charged
At high pH [H+] is low, therefore at a pH>pI proteins will be (-) charged
When pH=pI a protein stops moving

 

[Jo07]

this is taken from http://chimge.unil.ch/En/ph/1ph81.htm

Each amino acid has a specific isoelectric pH (pHi the pH at which amino acids are electrically neutral). At pH differing from pHi, amino acids bear an overall charge and therefore migrate under the effect of an electric field. Thus, working at a fixed pH (buffered solution) allows one to separate various amino acids by electrophoresis.

When an amino acid is placed in an electric field, it migrates towards the electrode of opposed polarity, while the neutral (zwitterionic) molecules do not migrate. Therefore :


When pH > pHi the amino acid bears a negative total charge: it migrates towards the positive electrode (anode).


When pH < pHi the amino acid bears a positive total charge: it migrates towards the negative electrode (cathode).


When pH = pHi the amino acid is in its zwitterionic, neutral form: it does not migrate and remains at the starting point.



I'm confused as to where the protein will migrate. On your post you say CAtion migrates to cathod and vice versa. I thought the positive electrode was the cathod, but this website says it is the anode.

Can someone clear this up for me?

 

[predentlsu1]

Nooo....There are two types of electrolytic cells. You have the Galvanic cells and the Electrolytic cells.
In Galvanic cells the anode is (-) and the cathode is (+)
In Electrolytic cells the anode is (+) and the cathode is (-)

Regardless of what type of cell, Reduction will always occur at the Cathode and Oxidation will always occur at the Anode (AN OX/RED CAT) Therefore cations such Na+ will always move toward the Cathode to get reduced (ie gain electrons)

Let me know if that helps.....

 

[predentlsu1]

There is a typo......there are two types of ELECTROCHEMICAL cells......not electrolytic cells sorry!!

 

[drteeth]

this is taken from http://chimge.unil.ch/En/ph/1ph81.htm

Each amino acid has a specific isoelectric pH (pHi the pH at which amino acids are electrically neutral). At pH differing from pHi, amino acids bear an overall charge and therefore migrate under the effect of an electric field. Thus, working at a fixed pH (buffered solution) allows one to separate various amino acids by electrophoresis.

When an amino acid is placed in an electric field, it migrates towards the electrode of opposed polarity, while the neutral (zwitterionic) molecules do not migrate. Therefore :


When pH > pHi the amino acid bears a negative total charge: it migrates towards the positive electrode (anode).


When pH < pHi the amino acid bears a positive total charge: it migrates towards the negative electrode (cathode).


When pH = pHi the amino acid is in its zwitterionic, neutral form: it does not migrate and remains at the starting point.



I'm confused as to where the protein will migrate. On your post you say CAtion migrates to cathod and vice versa. I thought the positive electrode was the cathod, but this website says it is the anode.

Can someone clear this up for me?

Remember that in an electric cell such as a Galvanic cell, oxidation occurs at the anode and reduction occurs at the cathode. Thus, electrons are lost at the anode leaving behind a positive charge and electrons are gained at the cathode building up a negative charge. Thus negatively charged anions go to the anode while positively charged cations migrate to the cathode. In the galvanic cell this is why salts are used to balance the charges that are built up... Hope this helps...

 

[Jo07]

Thanks guys