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Chromatography Columns and Consumables Catalog – Gas Chromatography Product Literature

What product do you recommend for removing sodium azide from the Alix Monoclonal Antibody (3A9) (Cat. No. MA1-83977) storage solution? Product FAQ

Answer

We recommend using Zeba Spin Desalting Columns, Plates, and Cartridges (Cat. No. 89882, 89883) to remove sodium azide from the Alix Monoclonal Antibody (3A9) storage solution.

Answer Id: E21291

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2019/2021 Chromatography Consumables Catalog - Liquid Chromatography Product Literature

What formats are available for the Melon Gel Support? Product FAQ

Answer

There are three different formats. The Melon Gel IgG Purification Kit (Cat. No. 45212) and Melon Gel IgG Spin Purification Kit (Cat. No. 45206) are for purifying antibodies from serum. The Melon Gel Monoclonal IgG Purification Kit (Cat. No. 45214) is for purifying monoclonal antibodies from cell-culture supernatant and ascites fluid. Ready-to-use prepacked chromatography cartridges are available as two 1 mL columns (Cat. No. 89932) or one 5 mL column (Cat. No. 89933).

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.

Answer Id: E8640

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What primer purification options are available, and how should I choose between them? Product FAQ

Answer

Invitrogen Custom DNA Primers are offered with several options for primer purification. Which one you should choose depends on your application. Here are some details on the options that are typically available:

Desalted--this is our standard purification procedure. The primer will be gas-phase deprotected and normal-phase desalted. Organic products are removed with 10% acetonitrile/90% water wash, and the products are then eluted under aqueous conditions. The smallest oligo we synthesize at this purification is 5 bases. Desalting removes salt, but not failure sequences. This method is sufficient for standard PCR, RT-PCR, cDNA synthesis, and sequencing reactions.

Cartridge Purification--this is based on reversed-phase chromatography to provide full-length sequences. The majority of the n-1, n-2, n-3, etc. species will be removed for oligos less than 35 bases. The smallest oligo we synthesize at this purification is 7 bases. Cartridge Purification is recommended for PCR with primers that have critical 5' sequences (RE sites, promoters), site-directed mutagenesis, first-strand cDNA synthesis for generation of libraries, gel shift assays, and production of cloning adapters.

HPLC (high-pressure liquid chromatography)--HPLC is a reversed-phase chromatography method similar to that used in cartridge purification. This process removes failure sequences and unincorporated labels. This is offered for oligos up to 50 bases long. HPLC is recommended for antisense work.

PAGE Purification--a denaturing polyacrylamide gel is used to purify the full-length primers from ALL types of n-1, n-2, etc. species of oligos. Oligos are purified by electroelution from the acrylamide gel and then run on a desalting column. These primers are not phenol extracted. PAGE purification can be used for any length of oligonucleotides. Typical yields for PAGE purified primers are not as high as when purified by other methods, but the product is highly pure. This is recommended for GeneTrapper screening. For more information, refer to the Oligos pages under Products & Services on our website.

Answer Id: E2933

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2019/2021 Chromatography Consumables Catalog - Sample Preparation Product Literature

What is the difference between HisPur Nickel Resin and HisPur Cobalt Resin? Product FAQ

Answer

Nickel has a higher affinity for histidines than does cobalt. It binds multiple histidines more tightly than cobalt, and requires more stringent conditions than is necessary for cobalt. The lower affinity of cobalt for multiple histidines typically results in less nonspecific binding of histidine-rich proteins that lack a his-tag compared to nickel resins.

HisPur Ni-NTA resin has a high capacity of up to 60 mg of 6xHis-tagged protein per milliliter. It is a versatile resin that can work under both native and denaturing conditions, and can also be used with a variety of lysis reagents and buffer additives.

HisPur Cobalt resin utilizes proprietary tetradentate chelating resin charged with cobalt. This system recovers highly purified protein with lower imidazole concentrations and has low metal leeching properties.

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.

Answer Id: E12933

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Can ProBond or Ni-NTA beads be used for large-scale preparations? Product FAQ

Answer

ProBond and Ni-NTA beads can be used in FPLC columns. However, the beads can only withstand low pressure (~43.5 psi max).

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.

Answer Id: E12988

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Are columns provided in the Melon Gel Purification kits? Product FAQ

Answer

Microcentrifuge spin cups are provided in the Melon Gel IgG Spin Purification Kit (Cat. No. 45206). No columns are included in the other Melon Gel Kits (Cat. No.s 45212 and 45214) so that researchers can adjust the scale of their purification. A wide variety of disposable columns are available for use with these kits. The product instructions for the Monoclonal Kit include a procedure for using a filter funnel and flask instead of a column for large-scale (e.g., 1 liter purifications.

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.

Answer Id: E8642

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After desalting with your Zeba desalting columns, I still have leftover salts in my sample. What should I do to get rid of all the salt? Product FAQ

Answer

You can either reduce the volume of sample processed in each column (the product manual indicates the volume range recommended for each column size) or run the flow-through from one column over a fresh column.

Answer Id: E12876

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What should the typical protein recovery be when using the Probond Purification System or Ni-NTA Purification system? Product FAQ

Answer

Both systems are qualified by purifying 2 mg of myoglobin protein on a column and performing a Bradford assay. Protein recovery must be 75% or higher.

Find additional tips, troubleshooting help, and resources within our Protein Purification and Isolation Support Center.

Answer Id: E12934

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How do your Zeba desalting products work? Product FAQ

Answer

A centrifuge is used to first remove the resin‘s void volume of liquid, followed by sample addition and centrifugation. After centrifugation, the macromolecules in the sample have moved through the column in approximately the same initial volume, but the small molecules have been forced into the pores of the resin and replaced by the buffer that was used to pre-equilibrate the gel-filtration matrix. Spin formats eliminate the need to wait for samples to emerge by gravity flow and require no chromatography system, allowing for multiple-sample processing simultaneously.

Note: The addition of larger volumes of buffer or longer centrifugation times than listed in the protocol will result in smaller molecules eventually emerging from the desalting column.

Find additional tips, troubleshooting help, and resources within our Protein Dialysis, Desalting, and Concentration Support Center.

Answer Id: E12863

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Are Zeba columns compatible with salts/organics? Product FAQ

Answer

Yes, Zeba columns are compatible with most salts. The resin is stable to some organics. As organics may affect performance, we suggest using less than 10% organics.

Answer Id: E12864

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Can I remove DyLight, Alexa Fluor or other dyes using the desalting columns? Product FAQ

Answer

No, excess amounts of DyLight dyes (and any other planar molecule with multiple rings in their structure) “act” larger than their stated molecular weights in desalting columns. We do not recommend desalting, but refer researchers to our Pierce Dye Removal Columns (Cat. No. 22858) for this purpose.

Answer Id: E12866

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Can these columns be regenerated and stored after use? Product FAQ

Answer

The desalting columns are designed to be used once and discarded after use.

Find additional tips, troubleshooting help, and resources within our Protein Dialysis, Desalting, and Concentration Support Center.

Answer Id: E12867

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