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Krypton Protein Stain FAQ


Answers to frequently asked questions (FAQs) about Krypton Fluorescent Protein Stain


 

What is the sensitivity of Krypton Protein Stain?
The stain is sensitive down to 0.25 ng using the standard protocol, or down to 2 ng using the rapid 30 minute protocol. A broad range of proteins have been tested with molecular weights from 14.4 to 200 K, including glycoproteins and phosphoproteins. Some of the proteins tested are as follows: myosin, beta-galactosidase, phosphorylase B, bovine serum albumin, ovalbumin, carbonic anhydrase, soybean trypsin inhibitor, lysozyme, aprotinin, horseradish peroxidase, myokinase, avidin, glucose oxidase, a2-macroglobulin, a1-acid glycoprotein, and beta-casein. (HeLa cell and rat heart tissue lysates were tested with 2D applications).

What gel types have been used with the Krypton Protein Stain?
Krypton Protein Stain has been used with Thermo Scientific Precise Protein Gels (Tris-HEPES), Invitrogen* Novex* Gels (Tris-glycine) and NuPAGE* (Bis-Tris) and Bio-Rad Criterion* Gels (Tris-HCl). Krypton Protein Stain is also compatible with homemade gels, native gels and 2-D gels. Sensitivity may vary depending on gel type.

What are the excitation/emission maxima and the extinction coefficient for the Krypton Protein Stain?
The excitation/emission maxima are 520 nm/580 nm with an extinction coefficient of 105,000 per mol cm.

How long does it take to stain gels using the Krypton Protein Stain?
The standard protocol takes only 2.5 hours to complete. A rapid, 30 minute protocol is also available that is sensitive down to 2 ng.

Why are so many proteins easily stained with the Krypton Protein Stain?
Ionic interactions cause the stain to bind basic amino acid residues, and hydrophobic interactions allow it to bind to alkyl chains. The dual-binding action makes the Krypton Protein Stain a universal stain for protein staining.

Does the Krypton Protein Stain produce a linear response for quantitative analysis?
Yes. The Krypton Protein Stain produces a linear quantitative range of three-to-four orders of magnitude.

How photostable is Krypton Protein Stain?
Krypton Protein Stain has good photostability. Typically, only an 8% decrease in signal occurs after 16 sequential scans.

Has the Krypton Protein Stain been compared to other stains?
Yes. Bio-Rad Flamingo*, GE Healthcare Deep Purple* and Invitrogen Sypro* Ruby Stains, all had problems not observed with Krypton Protein Stain. For example, Flamingo* Stain saturates and requires a full five hours for staining, Deep Purple* Stain produces fuzzy spots on 2-D gels and Sypro* Ruby Stain is expensive and the least sensitive of the four.

Does Krypton Protein Stain label nucleic acids?
No.

Is Krypton Protein Stain compatible with 2-D electrophoresis and mass spectrometry?
Yes. HeLa cell and rat heart tissue lysates separated by 2D electrophoresis and stained with the Krypton Protein Stain provided excellent sensitivity while not over-saturating highly concentrated spots. MALDI tryptic peptide mass fingerprinting of BSA and myoglobin resulted in characteristic profiles. The proteins were destained and processed with the Pierce In-Gel Tryptic Digestion Kit (Product # 89871). The lack of peak shifts indicates that the dye was not bound to the proteins after destaining.

How is the Krypton Protein Stain's signal captured?
Filter sets designed for Cy3 will work with the Krypton Protein Stain. Pierce scientists have used GE Healthcare Typhoon* 9410 Variable Mode Imager (532 nm laser excitation; 580 BP 30 emission filter), Kodak* Imager (CCD; 535 nm excitation and 600 nm emission filters), and visible light box. Sensitivity is better with lasers than with transilluminators.

What is the storage temperature for the Krypton Protein Stain and how stable is it?
Store Krypton Protein Stain at 4°C; although it is stable enough for ambient shipping. The 10X solution is under warranty for one year from the date of purchase if handled and stored properly. The Working Solution (1X) is stable at 4°C for 1-2 weeks.

If the stain is cloudy, can I still use it?
Yes. The solution will be cloudy immediately upon dilution to 1X, especially if it is cold. The cloudiness should dissipate with mixing.

 

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