Frequently Asked Questions

» Biotin-binding Proteins

» Glyko® Products

» FACE® Products

» Marker Enzymes

» PhycoPro™ Phycobiliproteins

» PhycoLink® Fluorescent Products

» Site-Specific Proteases

Biotin-binding Proteins

• Is Streptavidin with an activity of 16 U/mg from one company any different than Streptavidin at 15 U/mg from another?
• What is the sequence of Streptavidin?
• How stable is Streptavidin? How should I store it?
• I am using a Streptavidin agarose system to affinity purify biotin labeled, outer membrane proteins of a gram negative bacterium. Could you provide info regarding preparation of buffers and the reaction and elution conditions of this system?

Glyko® Products

• What happened to Glyko, Inc.? (letter describing purchase of Glyko by ProZyme, Inc.)
• I want to deglycosylate my antibody. Should I follow the denaturation protocol suggested in the Technical Data Sheet for N-Glycanase® (PNGase F)?
• What Glyko products do I need for glycan profiling of an antibody by HPLC using 2-AB fluorescent label?

FACE® Products

• ProZyme currently doesn't have N-Linked Oligosaccharide Preparative Gel listed as a product.  Is there an alternative?  Can I use an N-Linked FACE® Oligosaccharide Profiling Gel (GK60000) for preparative purposes?

Marker Enzymes

• What is the optimum temperature stability and pH stability of beta-galactosidase from E. coli?  Is the enzyme inhibited by either glucose or galactose?

PhycoPro™ Phycobiliproteins

• What does the R in R-Phycoerythrin stand for?
• What is the molecular weight of cross-linked allophycocyanin?

PhycoLink® Products     See also PhycoLink® TechNotes

» General

» Antibody Concentration

» Buffer/Formulation

» Calculating Concentrations

» FRET

General

• My conjugation didn't work. What do I do now?
• Why are phycobiliproteins so bright compared to other fluorescent dyes?
• How do I purify my conjugate to remove unreacted components?
• When I mix the reduced antibody solution with the SMCC-Allophycocyanin reagent, a precipitate forms.
• My protein is not an antibody and does not have any free sulfhydryls. How can I conjugate it to an SMCC-activated phycobiliprotein?
• I need more of the desalting columns (A & B) that are supplied with your kits.  Where can I get them?  Can they be re-used/regenerated?
• My conjugate gives me a high background.
• Can the PhycoLink Conjugation Kit be used to scale up to 10 mg of antibody?

Antibody Concentration

• My antibody concentration is less than 2 mg/mL. Must I concentrate it before performing a conjugation?
• In the instruction manual for the PhycoLink Conjugation kits, you suggest altering the reagent proportions for my specific antibody. What happens when I reduce the amount of antibody in relation to Allophycocyanin?
• The standard reaction size for your PhycoLink Allophycocyanin (PJ25K) and R-Phycoerythrin (PJ31K) Conjugation Kits is 1 mg of antibody. Can I conjugate four each of 0.25 mg of antibody instead? How about 10 each of 0.1 mg of antibody?
• I have recently used your R-Phycoerythrin and Allophycocyanin PhycoLink conjugation kits.  Am I right in assuming that one phycobiliprotein molecule binds one IgG molecule?

Buffer/Formulation

• My antibody is formulated in 50% glycerol. Must I exchange it into your suggested buffer?
• Why do you use pentochlorophenol?  Can I use another preservative such as azide?  Can I store your products without preservative?
• Does azide in the antibody formulation interfere with the conjugation protocols in the PhycoLink Kits?
• The antibody we wish to conjugate is currently in a lyophilized form.  Is there a preferred buffer we should use?

Calculating Concentrations

• How do I calculate the concentration of phycobiliprotein conjugates? (R-Phycoerythrin, Allophycocyanin and others)
• How do I calculate the molarity of phycobiliprotein conjugates? (R-Phycoerythrin, Allophycocyanin and others)
• How do I correct for blue dextran when determining the concentration of a phycobiliprotein conjugate? (R-Phycoerythrin, Allophycocyanin and others)

FRET

• Can phycobiliproteins be used in Fluorescent Resonance Energy Transfer (FRET)?
• In FRET assays, Europium is paired  with Allophycocyanin as the Donor/Acceptor pair.  Are other pairs available?
• I am using a cross-linked Allophycocyanin conjugate in a proximity assay (TR-FRET). If I increase the conjugate concentration, instead of getting an increase in signal, I get a decrease!

Site-Specific Proteases

• Factor Xa Purity - How pure is your Factor Xa?
• My fusion protein contains a thrombin cleavage site.  In the presence of Factor Xa, the protein is cleaved at the thrombin cleavage site as wells as the Xa site.  Is the Factor Xa contaminated with thrombin?