Technical FAQ

Technical FAQ

  1. Why are the proteins lyophilized? How will lyophilization affect the characteristics of the proteins?
  2. Why are protectants added to protein solutions before lyophilization? What protectants do you add to your products?
  3. How to reconstitute the lyophilized powder? Should I use sterile water, PBS or other buffer?
  4. Why are some proteins fused to tags?
  5. Do protein tags affect protein activity?
  6. Can Sino Biological Inc.'s products be used for other applications which are not listed on Sino Biological Inc. Certificate of Analysis?
  7. How should I store the products?
  8. How do you determine the quantity of your proteins and antibodies?
  9. Why is the quantification of the protein generated by my assay different from the one on the label?
  10. Who should I report to if I find the product quantity is incorrect? How will you deal with this kind of complaints?
  11. What materials are provided in your ELISA Kit?
  12. How many samples could be detected with each ELISA Kit?
  13. How should I store Sino Biological Inc.'s Antibodies?
  14. What is the shelf life of Sino Biological Inc.'s antibodies?
  15. How are Sino Biological Inc.'s Antibodies supplied?
  16. My Sino Biological Inc.'s antibody was shipped without ice but the recommended storage conditions are listed as +4°C or -20°C, will the activity have been affected?
  17. Can you provide me with the immunogen sequence for the antibody?
  18. The antibody I'm interested in isn't validated for the species I want. Will it work?
  19. Will Sino Biological Inc.'s antibody work in an application that is not listed in the applications table?
  20. Which dilutions of the antibody should I use in my experiment?
  21. What enzyme restriction sites were used to clone your cDNA into the vector and is it possible to reuse those sites to sub-clone?
  22. Is the cDNA cloned into the pMD18-T Simple Vector the same as the LacZ gene?
  23. How to obtain the cDNA sample from the filter paper?

1. Why are the proteins lyophilized? How will lyophilization affect the characteristics of the proteins?

Lyophilization (freeze-drying) technique is applied to increase the stability and shelf life of proteins, and decrease shipping cost. Lyophilization may cause loss of activity or aggregation of proteins. But addition of protectants (stabilizers, additives, excipients) and proper control of lyophilization conditions will minimize the adverse effect for most proteins.

2. Why are protectants added to protein solutions before lyophilization? What protectants do you add to your products?

Protectants (stabilizers) are added to protect proteins during lyophilization and/or long-term storage. Proteins are subjected to various stresses generated by lyophilization which may cause loss of activity, aggregation, or denaturation. Protectants can alleviate the stresses by several mechanisms including formation of an amorphous glassy state, replacing water, hydrogen bonding to proteins, physical dilution and separation of protein molecules, etc. Common protectants/stabilizers include sugars, polyols, polymers, surfactants, as well as some proteins and amino acids. We use trehalose and mannitol (normally 8% w/v) as protectants for lyophilization. Trehalose is a non-reducing disaccharide, well-known for its ability to stabilize biomolecules and microorganisms during prolonged period of desiccation. Mannitol is also commonly used as a stabilizer as well as a bulking agent in the process of lyophilization. It was reported to reduce aggregation for some proteins during lyophilization.

3. How to reconstitute the lyophilized powder? Should I use sterile water, PBS or other buffer?

We recommend using sterile water for reconstitution. Add the recommended volume of sterile water of to the vial, and gently shake it to solubilize the protein completely. Do not shake violently. PBS or other buffer can also be used as reconstitution agent as the salts in PBS can be omitted when the concentration and reconstitution volume is low. If possible, we suggest you compare the results of different reconstitution agents.

4. Why are some proteins fused to tags?

The initial use of protein tags is for protein purification purpose. In addition, tags are also used for protein detection in several applications, such as western blot and ELISA, when the specific antibodies are not available. Furthermore, Fc tag stabilizes molecules, which may increase the half-life of the linked products. Since the Fc fragment tends to dimerize, it helps the linked protein, particularly receptors, to form biologically active dimers. SBI also has human IgG1 Fc region (10702-HNAH ) and mouse IgG1 Fc region (10690-MNAH ) for purchase as a control.

5. Do protein tags affect protein activity?

It is different from case to case. For some applications, small tags, such as His-tag and FLAG-tag, may not affect protein activity and do not need to be removed. For example, there are more than 100 structures of His-tagged proteins in the Protein Data Bank. This indicates that the small His-tag often do not interfere with correct protein folding. Additionally, tested activity results are listed on our protein web pages, if you have concerns about tags interfering with protein activity. If there is no activity data online and activity is crucial to your experiments, please feel free to contact us (order@sinobiological.com) for latest information. For some recombinant proteins from SBI, fusion tags are removable by proteases. Please contact us for details.

6. Can Sino Biological Inc.'s products be used for other applications which are not listed on Sino Biological Inc. Certificate of Analysis?

If a specific application is not listed in SBI Certificate of Analysis and not tested by SBI yet, please send us your requirements to order@sinobiological.com. We will check if we could perform the assay or if any published data of the same product and the same application are available.

7. How should I store the products?

Lyophilized proteins are stable for at least twelve months from date of receipt when stored at -20℃ to -70℃. Upon reconstitution, the proteins can be stored at 2℃ - 8℃ for at least 1 month without detectable loss of activity. Reconstituted protein can also be aliquoted and stored frozen at -20℃ to -70℃in a manual defrost freezer for six months without detectable loss of activity. Avoid repeated freeze-thaw cycles. cDNAs can be stored at 2℃ - 8℃ for at least 6 months.

8. How do you determine the quantity of your proteins and antibodies?

Normally we determine the concentration of proteins and antibodies by measuring their UV absorbance. The concentration of proteins and antibodies is calculated by their extinction coefficients. We also use BCA, SDS-PAGE, HPLC and other methods to quantify the proteins and antibodies.

9. Why is the quantification of the protein generated by my assay different from the one on the label?

Different assays generate different quantification result. Sometimes the discrepancy can be significant if you conduct a different assay. It is also possible that the protein forms aggregations during storage which causes loss after reconstitution and centrifugation. We run quality control tests for each batch of proteins and antibodies. However, it is still inevitable that a few vials are not the same with the rest of the same batch. This happens very rare and can be minimized by our effort on Quality Control.

10. Who should I report to if I find the product quantity is incorrect? How will you deal with this kind of complaints?

You can call us at +86-400-890-9989 or send us an email to order@sinobiological.com. We will repeat your quantification assay on the same batch of product that was sent to you. If there is a discrepancy between the two assays, we will perform more types of assays to determine the quantity. If the quantity is actually incorrect, we will make up the shortage for free and give you credits for your future orders.

11. What materials are provided in your ELISA Kit?

Currently our ELISA kits contain the Capture Antibody, the Detection antibody, and the Standard (recombinant protein). Other buffer solutions and Substrate stock solutions required for the development of sandwich ELISAs need to be prepared by customers.

12. How many samples could be detected with each ELISA Kit?

Each kit contains sufficient materials to run ELISA assays on five 96-well plates (5 x 96T). If you have any other question, please see the product's Certificate of Analysis or contact us directly.

13. How should I store Sino Biological Inc.'s Antibodies?

Specific storage recommendations are listed on the Certificate of Analysis. We recommend that all antibodies be aliquoted into smaller aliquotes and stored at -20° C or below. Avoid multiple freeze-thaw cycles as this will affect antibody activity. We cannot guarantee how the performance of the antibodies if they are not stored as stated on the datasheet.

14. What is the shelf life of Sino Biological Inc.'s antibodies?

Unless otherwise indicated in the data sheet, most of our antibodies can be stored at 2℃-8℃ for one month without detectable loss of activity and are stable for twelve months from date of receipt when stored at -20℃ to-70℃. Repeated freezing and thawing should be avoided.

15. How are Sino Biological Inc.'s Antibodies supplied?

All Sino Biological Inc.'s antibodies are supplied as liquid. It remains stable during the delivery process at ambient temperature. We have tested every antibody's stability and activity following a period of more than 7 days at -20°C, room temperature, or 37°C. Antibodies under all three conditions turned out to be equally active and stable. For detailed information, please go to: http://www.sinobiological.com/Stability-of-our-antibody-a-1393.html

16. My Sino Biological Inc.'s antibody was shipped without ice but the recommended storage conditions are listed as +4°C or -20°C, will the activity have been affected?

No. Sino Biological Inc. ships thousands of antibodies every week to customers all over the world at ambient temperature. Loss of activity has never occured. We have tested every antibody's stability and activity following a period of more than 7 days at -20°C, room temperature, or 37°C. Antibodies under all three conditions turned out to be equally active and stable. For detailed information, please go to: http://www.sinobiological.com/Stability-of-our-antibody-a-1393.html However, upon receipt, antibodies should be stored according to the datasheet recommendations immediately.

17. Can you provide me with the immunogen sequence for the antibody?

The catalog number of the recombinant protein used as the immunogen is listed on the Certificate of Analysis. Please contact us via order@sinobiological.com if additional information is needed.

18. The antibody I'm interested in isn't validated for the species I want. Will it work?

Sino Biological Inc. cannot guarantee the performance of an antibody in an un-tested species. We can, however, make some educated guesses based the sequence that was used to generate the antibody. However, we cannot guarantee cross-reactivity even if the aligned sequences have a high identity score. We would greatly appreciate it if you would test our antibody on an un-validated species and share the results with us. We will issue you credits in return for your data.

19. Will Sino Biological Inc.'s antibody work in an application that is not listed in the applications table?

Customers can find all the tested applications for each particular antibody by visiting Sino Biological Inc. Antibody website. We do not list any un-tested applications. We would greatly appreciate it if you would test our antibody on an un-validated species and share the results with us. We will issue you credits in return for your data.

20. Which dilutions of the antibody should I use in my experiment?

We list a dilution suggestion on the datasheet for most of our antibodies. Please remember that these dilutions and concentration estimates are simply recommended as a starting point, and it may be necessary to adjust the dilution based on the experimental results.

21. What enzyme restriction sites were used to clone your cDNA into the vector and is it possible to reuse those sites to sub-clone?

The cDNA on pMD18-T Simple vector  is a TA clone, and multiple cloning site (MCS) is unavailable. Therefore, you would need to use specific PCR primers to sub-clone the inserted cDNA into other vectors. The cDNA on pGEM-T vector is a TA clone. But MCS is also available on this vector. Therefore, you could use either restriction enzyme digestion or PCR to sub-clone the inserted cDNA. The cDNA on  Vector 3 has MCS. The restriction enzymes used in each vector 3 clone and the maps is indicated in the products datasheet.

22. Is the cDNA cloned into the pMD18-T Simple Vector the same as the LacZ gene?

Since the pMD18-T Simple Vector is a TA-cloning vector, the cDNA insertion direction was not controlled. The inserts could be in either direction.

23. How to obtain the cDNA sample from the filter paper?

Please see the Usage Protocol below. Cut out the marked circle part from the center of the Whatman FTA elute card; Transfer this filter paper to a 1.5ml micro-centrifuge tube containing 150-200μl of sterile H2O. Make sure the filter paper is completely immersed in the sterile H2O. Incubate the tube at 95℃ for 15–30 min. After the incubation, pulse vortex or tap gently the tube approximately 60 times; Briefly centrifuge the tube for 30 seconds, and take out the supernatant. The supernatant contains the purified DNA. This DNA could be used for PCR or transformation directly.

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"