Electrophoresis Reagents

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Electrophoresis Reagents
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Electrophoresis

• Electrophoresis is a technique which separates
  charged biomolecules based on the rate at which
  they migrate in an applied electrical field.
• In many cases, electrophoresis of proteins are
  performed using polyacrylamide gel
  electrophoresis (PAGE). For molecular weight
  estimation and purity determination of proteins,
  sodium dodecyl sulfate (SDS)-PAGE is frequently
  employed.
• Agarose gel electrophoresis is one of the most
  common methods used to size-separate and analyze
  DNA. 

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Reagents for Gel Preparation, Buffer Preparation,
etc
A3217 30 Acrylamide / Bis-acrylamide (291) for Electrophoresis
A3218 30 Acrylamide / Bis-acrylamide (37.51) for Electrophoresis
A1132 Acrylamide Monomer for Electrophoresis
M0506 N,N'-Methylenebisacrylamide for Electrophoresis
A2098 Ammonium Peroxodisulfate for Electrophoresis
B3195 Bromophenol Blue Sodium Salt ( BPB) for Electrophoresis
D3647 DL-Dithiothreitol ( DL-DTT) for Electrophoresis
G0316 Glycerol for Electrophoresis
G0317 Glycine for Electrophoresis
M1948 2-Mercaptoethanol for Electrophoresis
S0588 Sodium Dodecyl Sulfate ( SDS) for Electrophoresis
T2515 N,N,N',N'-Tetramethylethylenediamine ( TEMED) for Electrophoresis
T2516 Tris(hydroxymethyl)aminomethane ( Tris-Base) for Electrophoresis
T3843 1-Thioglycerol for Electrophoresis
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Protein Electrophoresis- SDS

• SDS is a strong denaturant of proteins and is
  added to samples, gels, and buffer solutions for
  electrodes when proteins are separated with
  electrophoresis.
• As SDS not only denatures protein but also binds
  to the protein, when SDS is used in conjunction
  with a reducing reagent such as 2-mercaptoethanol
  to cleave disulfide bonds in the protein, and the
  protein is completely denatured, the amount of
  SDS bound is almost always proportional to the
  molecular weight of the protein.
• Resultantly, the protein is negatively charged.
  Therefore, the denatured protein can be separated
  by molecular weight independently of its
  structure and biological properties.

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Laemmlis Method - SDS

• Laemmlis method is the most widely used system
  of SDS-PAGE. In this method, the separation and
  the stacking gel contain Tris-HCl and the upper
  and lower buffer reservoirs contain Tris-glycine.
  All components of the system contain SDS.
• The advantage of Laemmlis method is that it
  gives sharper bands in the final plate.

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SDS-PAGE Sample Buffers

• Sample buffers are available in three different
  concentrations to allow for easy use with any
  sample volume. No reducing agent is included-add
  as required. A red sample bufferis also available
  to help prevent sample mix-up.
• For Example of use, Visit https//www.tcichemical
  s.com/IN/en/product/topics/electrophoresis_reagent
  s

B5834 2X SDS-PAGE Sample Buffer (2-Mercaptoethanol free) for Electrophoresis
B6104 4X SDS-PAGE Sample Buffer (2-Mercaptoethanol free) for Electrophoresis
B6105 6X Sample Buffer (2-Mercaptoethanol free) for Electrophoresis
B6110 2X SDS-PAGE Sample Buffer Phenol Red (2-Mercaptoethanol free) for Electrophoresis
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Gel Staining Reagent 

• Detecting protein bands in a polyacrylamide gel
  after electrophoresis requires the gel to be
  stained. Common staining methods include
  Coomassie Brilliant Blue staining, silver
  staining, fluorescence staining, and negative
  staining.
• Negative staining is a method in which only
  regions of gel containing protein remain
  unstained, while the remainder of the gel is
  stained white.
• Bands can be visualized by placing the gel
  against a dark background.

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Gel Negative-Staining Kit

• The Gel Negative-Staining Kit is used for Rapid
  and Highly Sensitive Protein Detection in as
  little as 20 minutes.
• Furthermore, stained gels can easily be destained
  using the included destaining solution, with the
  destained gel is able to be used in further
  downstream applications, such as Western blotting
  and mass spectrometry.
• G0615 Gel Negative Stain kit

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Example of use (Staining gels with G0615)

• 1) Place the post-SDS-PAGE gel in a tray
  containing enough deionized water to cover the
  gel and shake for 10 minutes.
• 2) Discard the deionized water, add enough
  solution A (diluted 10 times with deionized
  water) to cover the gel and shake for 5 minutes.
• 3) Submerge gel in deionized water for 10 seconds
  to wash. Repeat three times.
• 4) Transfer the gel to a new tray, add enough
  solution B (diluted 10 times with deionized
  water) to cover the gel and shake for 1 minute to
  develop color.

Figure. Photograph of gel stained by G0615
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Example of use (Preparation of gel for Western
Blotting)

• Place the stained and photographed gel in a tray
  containing solution C diluted 10-fold with
  deionized water.
• Shake the gel until the color is removed.
• Discard solution C, add enough deionized water to
  cover the gel and wash for 30 seconds. Repeat
  three times.
• Transfer the washed gel to a membrane (PVDF).

Figure. A Gels loaded with mouse serum and
stained with G0615.B The gel in A was destained
and mouse IgG was detected via Western Blotting.
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Gel Staining Reagent (Methanol-free CBB Stain
Solution)

• C3488 is a methanol- and acetic acid-free
  one-component ready-to-use solution for staining
  proteins.
• Example of use (Staining gels with C3488)
• 1) After gelelectrophoresis, wash the gel with
  deionized water for 5 minutes three times.
• 2) Remove the water, add C3488 till the gel is
  soaked and shake the gel gently for 1 hour at
  room temperature.
• 3) Remove C3488 and destain the gel with
  deionized water for 1 hour. If high background
  staining is observed, destain the gel with
  deionized water overnight at room temperature.

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Figure. Proteins stained by C3488 (destained
overnight)
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Reagents for Protein Staining and Others
A2097 Acid Black 1 ( Amido Black 10B) for Electrophoresis
A2256 Acid Red 112 ( Ponceau S) for Biochemical Research
B3193 Coomassie Brilliant Blue G-250 for Electrophoresis
B3194 Coomassie Brilliant Blue R-250 for Electrophoresis
F0718 Fast Green FCF for Biochemical Research
D1820 Sodium Deoxycholate for Electrophoresis
A2255 6-Aminohexanoic Acid for Biochemical Research
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DNA Electrophoresis

• The nucleotides that make up DNA carry a negative
  charge due to their phosphate groups, and are
  therefore attracted to the anode when run on an
  agarose gel.
• As the DNA moves through the gaps in the mesh of
  the agarose gel, longer molecules, with higher
  molecular weights, move more slowly, while
  smaller molecules are able to move more quickly.
• This method, of separating molecules by size on a
  gel using electricity, is known as gel
  electrophoresis.

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Nucleic Acid Staining Reagent (Ethidium Bromide
Solution)
Nucleic Acid Sample Preparation Reagents for
Electrophoresis
L0393 6X Loading Buffer Bromophenol Blue for Electrophoresis for Molecular Biology (1 mL3)
L0440 6X Loading Buffer Double BX for Electrophoresis for Molecular Biology (1 mL3)
Ethidium bromide is widely used as a DNA staining
agent with agarose gels and can detect DNA with
high sensitivity through visualization using UV
light. However, its carcinogenicity means it must
be handled with care. E1363 comes in a dropper
bottle pre-dissolved, making it safe and easy to
use. E1363 Ethidium Bromide (0.5mg/mL in
Water) (in Dropper Bottle) for Electrophoresis
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Example of use (Staining gels with E1363)

• 1) After electrophoresis, dilute E1363 (1 drop /
  40 mL) to 0.5 µg/mL with water or running buffer
  and incubate the gel for 15 minutes.
• 2) If you have to decrease background
  fluorescence, wash the gel in water for 15
  minutes.
• 3) In use of electrophoresis buffer solution,
  Ethidium bromide incorporated into nucleic acid
  and can visualize band immediately by using UV
  transilluminator.

Figure. DNA Ladder Marker stained
by E1363 (destained 15 minutes)
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Nucleic Acid Staining Reagent (Nucleic Acid Stain
Blue)

• Nucleic acid stain blue is used to stain nucleic
  acids after agarose gel electrophoresis. As
  nucleic acids are stained blue, no
  transilluminator or other detection device is
  required. Unlike ethidium bromide, it is
  non-mutagenic and therefore safe and easy to
  handle.
• N1209 10X Nucleic Acid Stain Blue
  for Electrophoresis

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Example of use (Staining gels with N1209)

• Prepare N1209 diluted 10 times with deionized
  water.
• After electrophoresis, immerse the gel in the
  diluted N1209 and shake for 10 minutes.
• Discard the staining solution and shake the gel
  with deionized water for 30 minutes and wash the
  gel. Repeat the wash step if background is high.

Figure. Photograph of gel stained
by N1209 (destained overnight)