Thermo Scientific™ PageRuler™ Unstained Low Range Protein Ladder

Catalog No.	Quantity
26-632-X4	8 x 250 μL
PI26632	2 x 250 μL

Catalog No. 26-632-X4 Supplier Thermo Scientific™ Supplier No. 26632X4

Description

Thermo Scientific PageRuler Unstained Low Range Protein Ladder is a mixture of eight purified proteins and peptides (3.4 to 100 kDa) for use as size standards in peptide and small-protein gel electrophoresis.

Thermo Scientific™ PageRuler Unstained Low Range Protein Ladder is a mixture of eight (8) purified proteins and peptides (3.4 to 100kDa) for use as size standards in peptide and small protein gel electrophoresis.

Thermo Scientific PageRuler Unstained Low Range Protein Ladder is a mixture of eight purified proteins and peptides (3.4 to 100 kDa) for use as size standards in peptide and small-protein gel electrophoresis. The protein ladder is supplied in a ready-to-use format for direct loading onto gels; no need to heat, reduce, or add sample buffer prior to use.

Product features

• Reference bands—25 kDa band is stronger than the others for easy orientation
• Tagged—the ladder proteins (except for the 5 and 3.4 kDa peptides) contains an integral Strep-tag™ II sequence and can be detected on western blots using Strep-Tactin™ conjugates

Applications

• Accurate protein sizing on SDS-polyacrylamide gels and western blots

Specifications

• Content And Storage: Includes eight vials of 250 μL each
  
  Storage buffer: 62.5 mM Tris-H₃PO₄ (pH 7.5 at 25°C), 1 mM EDTA, 2% SDS, 100 mM DTT, 1 mM NaN₃, 0.01% bromophenol blue and 33% glycerol
  
  Upon receipt, store at -20°C.
• Number of Markers: 8
• Ready to Load: Yes
• Size Range: 3.4 to 100 kDa
• Molecular Weight (g/mol): 100, 30, 25, 20, 15, 10, 5, 3.4 kDa
• Quantity: 8 x 250 μL
• Product Line: PageRuler
• Product Type: Protein Ladder
• Stain Type: Unstained
• System Type: SDS-PAGE

Frequently Asked Questions (FAQs)

I used one of your PageRuler unstained protein ladders and did not get good separation of the bands. What could have happened?

Here are possible causes and solutions:

- Ladder was boiled: Discard boiled aliquot.
- Too much volume of ladder used: Add less volume or dilute the ladder in protein loading buffer prior to use.
- DTT oxidation in storage buffer: Add freshly prepared DTT solution to a final concentration of 100 mM.

I used one of your protein standards for a western transfer and noticed that some of the lower-molecular weight protein bands passed through the membrane. How can I resolve this issue?

- Decrease voltage, current or length of transfer time
- Make sure that the methanol concentration in the transfer buffer is proper; use a methanol concentration of 10-20% methanol removes the SDS from SDS-protein complexes and improves the binding of protein to the membrane.
- Make sure that the SDS concentration (if added) in the transfer buffer is proper, don't use more than 0.02-0.04% SDS. Using too much SDS can prevent binding of proteins to the membrane.
- Check the pore size of the membrane and the size of the target protein. Proteins smaller than 10 kDa will easily pass through a 0.45 µm pore size membrane. If proteins smaller than 10 kDa are of interest, it would be better to use a 0.2 µm pore size membrane.

I used one of your protein standards for a western transfer and noticed that some of the higher-molecular weight bands transferred very poorly to the membrane. Can you offer some tips?

- Increase voltage, current or length of time for transfer
- SDS in the gel and in the SDS-protein complexes promotes elution of the protein from the gels but inhibits binding of the protein to membranes. This inhibition is higher for nitrocellulose than for PVDF. For proteins that are difficult to elute from the gel such as large molecular weight proteins, a small amount of SDS may be added to the transfer buffer to improve transfer. We recommend pre-equilibrating the gel in 2X Transfer buffer (without methanol) containing 0.02-0.04% SDS for 10 minutes before assembling the sandwich and then transferring using 1X transfer buffer containing 10% methanol and 0.01%SDS.
- Methanol removes the SDS from SDS-protein complexes and improves the binding of protein to the membrane, but has some negative effects on the gel itself, leading to a decrease in transfer efficiency. It may cause a reduction in pore size, precipitation of some proteins, and some basic proteins to become positively charged or neutral. Make sure that the methanol concentration in the transfer buffer is not more than 10-20% and that high-quality, analytical grade methanol is used.

I used one of your pre-stained standards on a Tris-Glycine gel and noticed that the molecular weights of the proteins were different than on a NuPAGE Bis-Tris gel. What is the reason for this?

Pre-stained standards have a dye that is covalently bound to each protein that will result in the standard migrating differently in different buffer systems (i.e., different gels). As a result, using a pre-stained standard for molecular weight estimation will only give the apparent molecular weight of the protein. Pre-stained standards may be used for molecular weight approximation, confirming gel migration and estimating blotting efficiency but for accurate molecular weight estimation, an unstained standard should be used.

I used one of your protein standards and am seeing some extra bands in the lane. Can you offer some suggestions?

- While loading, take care to make sure that there is no cross-contamination from adjacent sample lanes.
- Make sure that the correct amount of standard is loaded per lane. Loading too much protein can result in extra bands and this is a problem especially with silver-stained gels.
- Improper storage of the standard or repeated freeze/thawing can result in protein degradation.

I used one of your protein standards and the bands look non-distinct and smeary. What should I do?

Here are some suggestions:

- Make sure that the correct amount of standard is loaded per lane. Loading too much protein can cause smearing and this is a problem especially with silver stained gels.
- Bands will not be as well resolved in low percentage gels. Try using a higher percentage gel.
- If the bands look smeary and non-distinct after a western transfer/detection, this may be due to the antibody being too concentrated. Follow the manufacturer's recommended dilution or determine the optimal antibody concentration by dot-blotting.

A couple of bands in my protein standard are missing on the gel. Can you help me troubleshoot?

Here are some suggestions:

- Check the gel type/percentage of the gel that was used. Depending on the gel type and/or percentage, all the bands may not be seen. For example, the smallest bands of the protein standard may not resolve on a very low percentage gel whereas the higher molecular weight bands may not resolve on a high percentage gel.
- Check the expiration date on the protein standard. Expired lots may result in faded or missing bands due to protein degradation.
- Check the storage conditions for the protein standard. Improper storage conditions will compromise the stability of the proteins in the standard.
- Make sure that the protein standard was not heated/boiled prior to loading on the gel. Our protein standards are ready to load and we do not recommend heating/boiling them as this may cause degradation of proteins in the standard.

Are any animal-derived proteins present in the different PageRuler Unstained Protein Ladders?

The PageRuler Unstained Protein Ladder, PageRuler Unstained Broad Range Protein Ladder, and PageRuler Unstained Low Range Protein Ladder contain recombinant prokaryotic proteins and do not contain any animal-derived proteins.

Can the PageRuler Unstained Protein Ladder, PageRuler Unstained Broad Range Protein Ladder, and PageRuler Unstained Low Range Protein Ladder be detected using Strep-Tactin Conjugates?

The proteins in the different PageRuler Unstained Protein Ladders (except for the 3.4 and 5 kDa peptides in the PageRuler Unstained Low Range Protein Ladder and the PageRuler Unstained Protein Ladder, and the 5 kDa peptide in the PageRuler Unstained Broad Range Protein Ladder) contain an integral Strep-tag II Sequence and may be detected on western blots using Strep-Tactin Conjugates.

What is the composition of the PageRuler Unstained Low Range Protein Ladder?

The PageRuler Unstained Low Range Protein Ladder is a mixture of seven recombinant proteins ranging from 5 kDa to 100 kDa and a synthetic peptide at 3.4 kDa. For easy reference, the 25 kDa protein band is of greater intensity.

What are the storage conditions and shelf life for the PageRuler Unstained Protein Ladders?

We recommend storing the PageRuler Unstained Protein Ladders at -20 degrees C where they are stable for a year.

For Research Use Only. Not for use in diagnostic procedures.