HiMark™ Unstained High Molecular Weight

Protein Standard
Cat. no. LC5688 Size 250 µL Store at −30°C to −10°C
Pub. Part no. LC5688.pps MAN0001520 Rev. Date: 5 December 2011
The HiMark

Unstained High Molecular Weight (HMW) Protein Standard
allows you to accurately estimate the molecular weight of high molecular
weight proteins on NuPAGE
Novex Tris-Acetate Gels with Tris-Acetate
SDS buffer system.
The important features of the standard are:
• Consists of 9 protein bands in the range of 40–500 kDa
• Designed for use with NuPAGE
Novex 3-8% and 7% Tris-Acetate Gels
under denaturing conditions
• Supplied in a ready-to-use format
• Visualized with Coomassie or silver staining
• Visualized also with Ponceau S, Coomassie, or other membrane stains
after western transfer
Contents: 250 μL of HiMark

Unstained HMW Protein Standard
Storage 250 mM Tris-HCl, pH 8.5; 0.5 mM EDTA; 50 mM DTT;
Buffer: 10% glycerol; 2% LDS; 0.2 mM Coomassie G-250;
0.175 mM Phenol red
Storage: Store at −30°C to −10°C. Avoid repeated freezing and thawing
Stability: 6 months at −20°C
Product Use For research use only. Not for human or animal therapeutic or diagnostic
The HiMark

Unstained HMW Protein Standard is supplied in a ready-to-
use format. There is no need to heat or add reducing agent.
1. Thaw the standard at room temperature. Vortex gently to ensure the
solution is homogenous.
Note: If there is a precipitate in the standard, thaw at room temperature
for 10–15 minutes and vortex to solubilize or warm the solution at 30°C
(Do not heat at >37°C).
2. Load the ladder using the loading volumes listed below to obtain the
best results. Use an appropriate percentage of NuPAGE
Tris-Acetate Gel to resolve the proteins.
Gel Electrophoresis/Blotting
Mini-Gel (1.0-mm thick) 5 μL
Mini-Gel (1.5-mm thick) 7 μL
3. After electrophoresis, we recommend staining the standard proteins with
Silver Staining Kit, SimplyBlue

SafeStain, or Coomassie
stain. Avoid using the SilverXpress
Silver Staining Kit.
Blotting: To obtain higher transfer efficiency during the transfer of high
molecular weight proteins, avoid using methanol in the transfer buffer.
After transfer, you may stain the standard proteins on the membrane with
Ponceau S, Coomassie, or any membrane stain of choice.
Note: The HiMark

Unstained HMW Protein Standard is designed for use
with NuPAGE
Novex Tris-Acetate Gels. Using the standards with NuPAGE

Novex Bis-Tris Gels or Tris-Glycine Gels may result in inaccurate molecular
weight estimation.
Product Qualification
The Certificate of Analysis (CofA) provides detailed quality control
information for each product. The CofA is available on our website at
www.lifetechnologies.com/support, and is searchable by product lot
number, which is printed on each box.


Molecular Weight Calculator
A molecular weight calculator is available at www.lifetechnologies.com.
From the home page, go to Products & Services Controls > Standards, &
Ladders > Protein Standards & Ladders > Unstained Protein Standards.
Select HiMark

Standard and follow instructions on the page to download
the calculator.
The calculator allows you to easily and accurately calculate the molecular
weight of your proteins on NuPAGE
Novex 3–8% and 7% Tris-Acetate Gels
and to extrapolate the molecular weight of proteins beyond the standard

Unstained HMW Protein Standard (5 μL) was analyzed on a
Novex 3–8% Tris-Acetate Gel (A) or a NuPAGE
Novex 7% Tris-
Acetate Gel (B) and stained with SimplyBlue

SafeStain. The apparent
molecular weights of protein standard bands are shown below.

Accessory Products
The following products are available separately and may be used with the

Unstained HMW Protein Standard.
Quantity Cat. no. Product
Novex 3–8% Tris-Acetate Gels 1 box EA0375BOX
Novex 7% Tris-Acetate Gels 1 box EA0355BOX
Tris-Acetate SDS Running
Buffer (20X)
500 mL LA0041
LDS Sample Buffer (4X) 10 mL NP0007

SafeStain 1 L LC6060

Silver Staining Kit 1 kit LC6070

Limited Use Label License: Research Use Only
The purchase of this product conveys to the purchaser the limited, non-
transferable right to use the purchased amount of the product only to perform
internal research for the sole benefit of the purchaser. No right to resell this
product or any of its components is conveyed expressly, by implication, or by
estoppel. This product is for internal research purposes only and is not for use
in commercial applications of any kind, including, without limitation, quality
control and commercial services such as reporting the results of purchaser’s
activities for a fee or other form of consideration. For information on
obtaining additional rights, please contact outlicensing@lifetech.com or Out
Licensing, Life Technologies, 5791 Van Allen Way, Carlsbad, California 92008.
©2011 Life Technologies Corporation. All rights reserved. The trademarks
mentioned herein are the property of Life Technologies Corporation or their
respective owners.
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