Product Number CM9xxxx
CellMosaic’s product number (catalog number or SKU) CM9xxxx is generally assigned to any bioanalysis and bioassay related reagents and kits such as analysis standards.
CM90005: Thiol Assay Kit with Purification
What is the difference between CM90004 and CM90005?
CM90004 (thiol assay kit) is for checking purified biopolymer containing a thiol group. CM90005 (thiol assay kit with purification) is for assaying thiol groups after biopolymer modification. This kit includes a purification spin column to remove unreacted reducing reagents or thiolation reagents before the assay.
CM92004: SEC (Gel Filtration) HPLC Protein Standard (7 components, lyophilized)
How much total protein is in each vial of the SEC Protein Standard?
The products contain a mixture of proteins in a lyophilized format. Once you dissolve the product in 80 µL of deionized water, the concentration of each protein is within 0.01-1 mg/mL. There are approximately 1.5-3 mg of total protein content in one package (4x80 µL). We cannot supply the exact concentration of each protein because the product is designed for MW calibration and not for quantitation. If you need to perform a quantitation by HPLC, we suggest using BSA to create a standard curve.
Can I scale-up the recommended injection volume to match the length of my column?
Yes, the concentration of each indicator is optimized to use 8 μL for a standard SEC HPLC analytical column with a volume of 10-15 mL. If you have a larger volume column, you can scale the injection volume linearly. For example, if you have 200 mL resin, you may need to inject around 200 μL total (around 3 tubes, 4x80 μL provided).
If you are using a loop size that accommodates significantly more volume than the volume you need to inject, you will need to inject enough volume so that when the standard dilutes into the loop volume it will still be around 0.1-1 mg/mL for each protein.
Can I use CM92004 with a semi-preparative column?
For purification columns we generally suggest using our BSA standard for protein quantitation (CM92006), CM92004 is designed to be used in an analytical or gel filtration column. When used with columns packed using preparative resin, this standard will give broad, not well resolved, peaks and you will need to scale-up the volume of sample you are injecting (see “Can I scale-up the recommended injection volume to match the length of my column?” for more information).
Can I use CM92004 with an FPLC column (Cytiva column)?
Yes, you can use it with an FPLC column (Cytiva column). However, keep in mind that the peaks may not be as sharp as with the HPLC column shown in the product sheet, and the first two peaks (blue dextran and thyroglobulin) are not separable.
Our sample is a 1200 KDa protein. Can I use CM92004 to generate a standard curve to calculate the MW?
On Page 4 of the CM92004 product sheet, there is a representative Log MW vs. Rt(min) graph (Figure 2). As you can see, Dextran is not very representative of proteins. To obtain reliable data, you can use data points from other proteins (or maybe only the high MW ones) to generate a graph for the calculation.
I used CM92004 on a new Superdex 200 16/26 column (120 mL total volume). I injected 80 µL of the standard into a 2 mL loop, but I didn’t see any peak on the profile.
This SEC protein standard is designed to be used in an analytical HPLC or gel filtration column. Gel filtration columns using Superdex 200 will give slightly broader peaks, with peaks 1 and 2 being unresolved. The column you are using is a semi-preparative column (120 mL), which requires more sample. I saw you are trying to scale-up the amount based on the HPLC column. However, you injected 80 microliters of the sample into a 2 mL loop. The solution will be diluted within 2 mL before it can reach your column so you will probably not see anything besides some baseline increase. If you want to use this standard, you may need to change the loop size to 100 µL or inject more volume of the same concentration, which has around 0.1-1 mg/ mL of each protein. See if this works. But keep in mind, Superdex 200 preparative resin (large bead size) does not have good resolution for these proteins compared to the standard HPLC SEC resin. Usually for a purification column, we just use BSA as the standard. We do have a BSA standard (Cat# CM92006) you can purchase.
How do I improve the peak resolution and separation of the standard proteins for a particular column?
The separation in SEC is mainly determined by the resin/column you are using (particle size, modification, type). However, there are a few things you can do to improve the peak resolution and achieve better separation of the protein standards. For example, decrease the flow rate, decrease the injection loop size (not just the injection volume), shorten the line between the injector to the column, and clean the column before usage. The performance of the column decreases dramatically with use. Try to clean it and follow the manufacturer’s instructions before injecting the standard.
CM92005: SEC (Gel Filtration) HPLC Low Molecular Weight (LMW) Protein Standard
Can I scale-up the recommended injection volume to match the length of my column? (the protocol given with CM92005 is for a 10-15 ml column)
You can scale up to modify the protocol for your column. However, keep in mind that this product is recommended for use in HPLC columns. If you are using other, non-silica-based SEC columns, your peak resolution may not be as good, and the first two peaks may not resolve. If the signal is not good, you can continue to increase the amount injected.
What is the concentration of proteins provided in the standard?
This product contains a mixture of proteins in a lyophilized format. Once the product is dissolved in 80 microliters of DI water (according to the product sheet), the concentration of each protein is within the 0.01-1 mg/mL.