Services

Custom Polyclonal Antibody

FAQ

1. What kind(s) of antigen(s) can be injected?

Native proteins purified from the natural source, recombinant proteins  fragments or full length proteins (expressed in a variety of hosts such as E. coli, Mammalian cells  etc),

Chemical peptides (up to 22 aa)  or other compounds of microbial, fungal or virus origin.

If the antibodies are going to be used on the denatured target protein (example: Western blot, immunohistochemistry on fixed tissues), denatured forms of antigens are preferred (like gel piece, protein in inclusion bodies).

If the antibodies are going to be used on native target proteins (example: immunoprecipitation), non-denatured forms of antigen are preferred (protein in solution free of  denaturing agents).

2. How much antigen is required for immunization?

It usually depends on the immunogenicity of the antigen. In the standard protocol here are the quantities we usually need (per animal):

Rabbit : >1 mg of protein or >2.5 mg of conjugated peptide and 0.5mg of unconjugated peptide for ELISA.
Chicken : >2.5 mg of protein or >2.5 mg of conjugated peptide and 0.5mg of unconjugated peptide for ELISA.
Guinea Pig : >0.7 mg of protein or >1.5 mg of conjugated peptide and 0.5mg of unconjugated peptide for ELISA.
Rat : >0.7 mg of protein or >1.5 mg of conjugated peptide and 0.5mg of unconjugated peptide for ELISA.
Mouse : >0.4 mg of protein or >0.6 mg of conjugated peptide and 0.5mg of unconjugated peptide for ELISA.
Goat & Sheep : >10 mg of protein or >10 mg of conjugated peptide and 0.5mg of unconjugated peptide for ELISA testing.

3. Do I need to conjugate my immunogen?

Conjugation makes the immunogen more immunogenic by making it physically large enough for the immune system to recognize it.

We conjugate all the peptides we synthesize usually using keyhole limpet hemocyanin (KLH). Other good carrier proteins are bovine serum albumin (BSA) and ovalbumin (OVA).

We also recommend small proteins (less than 15KD) to be conjugated.

4. How much purified pAbs or antisera will I receive with a standard package?

It depends on the host species you chose.

Rabbit : 30-40 ml of antiserum or 1 -10 mg of purified antibodies.
Chicken : 30-40 ml of antiserum/project or 1 -10 mg of purified antibodies.
Guinea Pig : 2.5-5 ml of antiserum or 1-10 mg of purified antibodies.
Rat : 2.5-5 ml of antiserum or 1-10 mg of purified antibodies.
Mouse : 300-500µl of antiserum or 10-100 µg of purified antibodies.
Goat & Sheep : 500-1000 ml of antiserum or >100mg of purified antibodies.
Llamas/Alpagas : up to 800ml of serum or up to 750ml of serum and 50ml serum purified

5. How many animals should I use?

We suggest you to use at least 2 animals per project (except for llama/alpacas, where usually we only need one animal). There is often variability of immune response between individual animals, so this will increase your chances of getting a highly reactive and specific antibody to your protein of interest.

6. Are the final antibodies purified from the antisera pooled together from the two animals?

At ProteoGenix we usually do not pool the sera, so you will receive 2 different antibodies at the end of your project if you chose to immunize 2 animals. If you want us to pool the sera and/or antibodies, feel free to ask us !

7. Do you charge any fee if you fail to deliver pAb at your guaranteed titer?

Usually, when the titers do not reach our guarantee, we will try to discuss with you and understand what could have happen.

Then we will usually offer 2 possibilities:
-Restart the project free of charge until the guarantees are met.
-Cancel the project. In that case, only the set-up fees noted in your quotation will be invoiced.

Proteine Custom Service

Production & purification of recombinant proteins

Recombinant Protein Services

5 Expression systems allow us to cover all customer’s requirements:

Typical timining for recombinant protein expression

FAQ

1. How much protein quantities can I expect from 1L culture?

Recovery yield can vary a lot from one expression system to another and depends a lot on the protein to be produced.

Small scale expression and purification tests included into all our standard packages are especially intended to evaluate the recovery yield we can expect from 1L culture as requested. At the end of this step, you are informed about this through a detailed report and are free to decide to Go or No Go with the 1L culture production (or adjust the culture volume accordingly).

2. This protein is easy to produce, I want to do directly 1L production

Even if the customer has the feeling that the protein is easy to produce (or even if he has already produced the protein before), we always have to recommend him to do the 1st step of small scale expression/purification evaluation for the following reasons:

-The customer doesn’t say the true and/or may underestimate the difficulty. We have therefore to check before to go to 1L production to avoid any complains.
– Results and experiments described into the literature are often challenging to reproduce (up to 70% of results described in papers can’t be reproduced according to recent studies).
– Variations can occur from one lab to another one.

3. Can we send you our expression vector for the production

Yes it’s possible. We just need to receive the vector map and full sequence of the expression vector for validation by our technical team before to be able to make a quotation.

Extra cost may be required in case of expensive antibiotic selection, subcloning into another vector…

It is important to ask if the cDNA was optimized for the host expression system and to check this. We always recommend to optimize the cDNA to ensure the best results. If the cDNA provided by the customer is not optimized, you must suggest to customer a new gene synthesis with codons optimization.

4. What happen if you the protein can’t be expressed? What do you charge if the protein expression fails? 

You need to explain that work can be stopped at any time and only started or completed steps are invoiced and due.

For example, if we did the expression and purification tests and the protein can’t be expressed, we only charge these expression and purifications tests.

5. How are the payment conditions?

We usually ask for full prepayment of gene synthesis and subcloning because these steps will be done anyway.

About standard protein production packages, we ask for a 50% prepayment in order to cover at least the cost of small scale expression/purification tests.

Gene Synthesis / Subcloning / Mutagenesis

Gene synthesis workflow

Gene Synthesis

  Deliverable and guarantees

• Free codon optimization : the optimization will be sent to the customer after having received the order.
• We will not start the synthesis before we receive the customer’s validation of the final optimized sequence.
• Synthesis of the double Stranded cDNA
• Sequencing included
• Efficiency > 99.9% (basically, one error every 1000 base pair)
• Mutagenesis run to remove errors if needed
• Delivered sequence = 100% accuracy
• Shipment of ~4μg of the cDNA lyophilized and cloned in a vector
• High throughput service: synthesis of several genes in parallel without any additional time needed and with special discounts

FAQ

1.The gene synthesis failed, What do I pay?

No gene, no fee.

2. On what is based your codon optimization tool?

We take 4 aspects into account:
-CAI value (corresponds to codon usage table on the organism)
-GC content
-Repetitions
-Restriction sites
-Secondary structures

3. How do you guarantee the sequence accuracy of my genes? What QC data will be provided? What’s the purity?

We provide 100% sequence-guaranteed synthetic genes.

If needed/upon request we can send complete QC data containing: construct map, sequencing chromatograms and certificate of analysis (COA).

The purity is based on the A280/A260 ratio. If this ratio is comprised between 1.8-2 we conclude that the DNA is pure

4. How are genes shipped? 

We deliver 4ug of lyophilized plasmid, which contains your target gene subcloned in our standard vector or in any vector of your choice.

We can prepare more plasmid upon request (see DNA preps).
Plasmids are shipped at room temperature.

5. What is the typical turnaround time for gene synthesis?

Peptide Synthesis

• +20 years of experience in this field; 99% success rate
• Quality comittment – we guarantee:
  • the amount
  • the purity from crude up to 99% (controlled by HPLC) and
  • the sequence of the peptide (validated by MS)
  • COA, HPLC and MS controls are provided
• Able to synthesize long and difficult peptides (peptides with repetitions, circular peptides, or highly hydrophobic for example)
• Able to provide large quantity of peptides (from 1 mg up to grams)
• Large range of possible modifications
• our large volumes of production allows us to lower the cost of production to the minimum.
• Peptide library synthesis and design
• Fmoc solid phase synthesis from Cter to Nter

Peptide modifications & conjugations

N/C- TERMINAL MODIFICATIONS