FAQ

The ingredient of # R0011 is not open. # R0011 does NOT contain ethidium bromide. The extracted RNA from polyacrylamide gel stained with RNA Staining Solution maintains the integrity (I mean no degradation is observed) and cloning efficiency of cDNA prepared from the extracted RNA is good.

The Small RNA Marker (#R0007) contains approximately 6.4 micro gram of RNA, which is a mixture of five single-stranded RNAs, 20, 30, 40, 50 and 100 bases. Roughly speaking, concentration of each fragment is approximately 0.05 mg/ml (concentration of each fragment is slightly different). If you load one or two micro L of Small RNA Marker (#R0007) to a well of polyacrylamide gel on electrophoresis, RNA bands are seen by ethidium bromide stain. In this case, you do not have to label the marker with radioisotope.

The Total RNA included in the one kit of Small RNA Marker (R0007) is approximately 6.4 micro grams, thus the concentration is approximately 0.2 mg/ml. The Small RNA Marker (R0007) has five single-stranded RNAs, 20, 30, 40, 50 and 100 bases. Concentration of each fragment is slightly different because each RNA amount was adjusted to get good visibility on electrophoresis. Be sure to that this RNA marker is not intended for use in quantitative analysis.

The system has been successful expressing proteins > 120 kDa (ie SARS S protein). While the yield may not be as high as proteins with MW <80Kd, we are able to compensate for this by using more lysates to produce the overall required quantity of protein for the customer.

A novel highly efficient and robust wheat germ cell-free protein synthesis system was developed by Professor Yaeta Endo of Ehime University, Japan. The RNA N-glycosidase tritin and other inhibitors such as thionin, ribonucleases, deoxyribonucleases, and proteases that originate from the endosperm and inhibit translation were removed by extensive washing of wheat germ embryo. The in vitro expression using the extract from wheat germ devoid of the inhibitors and optimized expression vector is highly efficient and stable, and is capable of translating a full length protein as long as 70-80 kDa. Publications related to Wheat Germ in vitro expression system are listed below:

1: Sawasaki T, Ogasawara T, Morishita R, Endo Y. A cell-free protein synthesis system for high-throughput proteomics. Proc Natl Acad Sci U S A. 2002 Nov 12;99(23):14652-7. Epub 2002 Oct 30. PMID: 12409616 [PubMed - indexed for MEDLINE]

2: Sawasaki T, Hasegawa Y, Tsuchimochi M, Kamura N, Ogasawara T, Kuroita T, Endo Y. A bilayer cell-free protein synthesis system for high-throughput screening of gene products. FEBS Lett. 2002 Mar 6;514(1):102-5. PMID: 11904190 [PubMed - indexed for MEDLINE]

3: Morita EH, Sawasaki T, Tanaka R, Endo Y, Kohno T. A wheat germ cell-free system is a novel way to screen protein folding and function. Protein Sci. 2003 Jun;12(6):1216-21. PMID: 12761392 [PubMed - in process]

4: Kigawa T, Yabuki T, Yoshida Y, Tsutsui M, Ito Y, Shibata T, Yokoyama S. Cell-free production and stable-isotope labeling of milligram quantities of proteins. FEBS Lett. 1999 Jan 8;442(1):15-9. PMID: 9923595 [PubMed - indexed for MEDLINE]

5: Madin K, Sawasaki T, Ogasawara T, Endo Y. A highly efficient and robust cell-free protein synthesis system prepared from wheat embryos: plants apparently contain a suicide system directed at ribosomes. Proc Natl Acad Sci U S A. 2000 Jan 18;97(2):559-64. PMID: 10639118 [PubMed - indexed for MEDLINE]

We use a very immunogenic strain of mice which is not Balb/c for monoclonal antibody production. We do not work on hamster and rat.

We screen hybridomas with ELISA and Western blot against provided protein. If you want other performance, you need to send us materials. We will test for you.

We will try our best to dissolve the insoluble including using 1.5% of sarcosin. We may also try to inject the insoluble protein retained in bacterial inclusion body to make antibody.

At least 20 amino acids.

If mice were immunized with conjugated peptide, we only guaranty to produce antibodies which recognize synthesized peptide, and we do not guarantee on the recognition of the peptide sequence in its native protein since synthesized peptide may not have the correct folding structure.

We make mostly IgG2a and IgG2b antibody.

Yes, it is an optional service. Abnova cannot guaranty that antibody will recognize the lysate since (1) there are uncertainties on the quality of lysates received from customers, and (2) antigens expressed in bacteria may have totally different conformation as protein expressed in mammalian lysates.

Yes. If cDNA or plasmid DNA were available, Abnova can provide vectors for cloning free of charge; alternatively, subcloning by Abnova is available with additional US$ 200.

Yes, customers receive all hybridoma clones which customers ask to develop. Customers will receive up to 5 clones under standard custom production service. Additional clones will be charged accordingly.

If you supply us with cDNA, we may sub-clone it into our vector, and express it with wheat germ in vitro protein expression system.

If the antigen is a peptide, 2 mg will usually be sufficient before conjugation (1 mg for KLH, and 1 mg for BSA). If the antigen is a protein, 500µg/mice at concentration of 0.5-1 mg/ml is usually sufficient.

Abnova's immunization protocols can be adapted for native proteins, recombinant proteins, and peptides. Antigens harmful to human health are not accepted.

Abnova uses non-fusion antibody library derived from entire immune repertoire instead of myeloma fusion (<1% fusion rate) for Mab generation and screening.

Rabbit Mab has broader antibody diversity, antigenicity, affinity, specificity, and applicability than mouse Mab.

It depends on the size (e.g., peptide vs. protein) and sequence homology (e.g., human vs. rabbit) of the antigen.

Customer

Antibody production charge is based on milestones, hence minimizes the risk to the customer.

Yes.

Yes.

Yes, including different species Fc, isotype Fc, biotin, streptavidin, HRP.

No. The plasmids are Abnova’s proprietary technology.

Yes. There is an extra charge for the antibody sequence but it can be decreased or waived depending on the scale of follow-up recombinant antibody production.

Yes.

Human S100B ELISA kit measures total S100B in serum, heparin plasma and cerebrospinal fluid samples. It is specific for S100 beta subunit, S100 beta-beta and beta-alpha dimers. It doesn´t crosreact with S100 A1, S100 P and S100 Z proteins.

I am afraid we did not test the cross-reactivity of rat samples in our Human Clara Cell Protein ELISA. It may work but we have no proof for it. We are going to accomplish cross-reactivity data in many assays including Clara Cell Protein ELISA soon.

Serum and heparin plasma are preferred for this assay. We cannot recomend to use EDTA or citrate plasma samples.

As you customer says, our labeling method is not via aldehyde. For your reference, below is our labeling method.

Moreover, conjugation efficiency is almost 100% and conjugate collection rate is more than 90％.

We usually obtain 5ml of ascites from each Balb/c mouse and 8-10ml from each SCID mouse. The custom ascites service is based on number of mice used to generate the quantity of ascites needed by the customer. However, we cannot guarantee the same yield for every cell line since each may have a distinct growth characteristic. However, if an absolute quantity is desired, we can fulfill the request by inject more mice.

Abnova requires the customers to provide medium information and growth rate of their hybridoma cell lines. If the customers have the isotyping QC data for the hybridoma, we would also require this information. On the other hand, we can perform isotyping test for the customer upon request.

Endogenous, un-transfected lysate is used as control sample for comparing generated antibody’s reactivity against the target protein on western blot. It is difficult to know the exact quantity of protein expressed by the un-transfected cells versus transfected cells a priori. Nevertheless, transfected cells tend to express a larger amount of protein of interest, which is more amenable to antibody detection. The western blot images of antibody against un-transfected and transfected cell lysates are taken in tandem, while considering factors such as primary and secondary antibody concentration, developing and exposure time interval, as to achieve the best outcome and comparative result.

Details of the payment will be advised on the invoice that will besent together with your shipment. The following methods of payment are accepted:
• Credit card
• Cheques
• Wire transfer
We do not accept money orders or cash.

If you find any defect upon receipt of the product(s), please inform us within 10 days. We will replace the faulty product(s) immediately. If you received a product that does not work in your experiment, you do not need to return it; however, we do need your experimental data for the next process. Please obtain a troubleshooting form from us to fill in; we will make sure that you receive full support from our professional team.