Production and Purification of Antibodies and other Proteins
We produce and purify proteins from culture supernatants. Cells can be grown either in roller bottles, CellLine Flasks or in a Hollow Fiber Bioreactor. Please click on the ‘Large-scale Mammalian or Insect Cell Culture’ tab or the ‘Transfection’ tab.
Antibody conjugation and Fragmentation
The facility can generate Fab fragments from purified monoclonal antibodies. The antibodies are digested by Papain overnight fragments are purified over a Protein A column.
Antibodies can also be conjugated to FITC or biotin.
Storage of customer cells in CCSF LN2 freezers
For a charge we will store customer cell vials in our LN2 freezers.
Growth of seed cultures for many common cell lines
Please email Sabine Baxter with your cell line request and we can check to make sure we have the cell line. Cells are mycoplasma tested and can be provided as live cells in a T25, T75 or T150 flask.
Mycoplasma Testing
Mycoplasma contamination cannot be detected with a light microscope. Cells have to be tested for contamination. It is recommended to test all cells in the lab at least every 6 months.
Culture your cells for at least two days without any antibiotics (without Penn/Strep, G418, etc.), prior to removing a sample cell culture supernatant for testing. Most common strains of mycoplasma grow slowly in the presence of antibiotics and may be present only in spurious amounts and thus may be undetectable. Testing cells grown with antibiotic can lead to false negative results because most mycoplasma strains are dormant but not dead in the presence of antibiotics.
We only need about 1ml of cell culture supernatant for testing exclusively in 1.5ml Eppendorf tubes. Please remove any cells that might be present by centrifugation. We run a mycoplasma test every Tuesday and Friday morning. Please note that samples should arrive at our facility by Monday or Thursday afternoon before 4:30 pm to be included in next day’s testing. You will receive results on Tuesday or Friday afternoon.
Detection of Mycoplasma by MycoAlert (Cambrex)
This test is extremely rapid, highly sensitive and is generic for all common mycoplasma strains. Mycoplasma have unique metabolic processes that derive energy from their host cells. The method specifically detects enzymes involved in mycoplasma metabolism that are absent in host cells. By using substrates for the mycoplasma specific enzymes, ADP is converted to ATP. The resultant ATP is conveniently detected using the bioluminescent reaction catalyzed by the Firefly Luciferase enzyme. Any increase in light output from this reaction, as measured by a luminometer, will indicate the presence of mycoplasma.
Please be aware that if your cells express luciferase, this will lead to a false positive result. A different kind of assay should be used for testing these cells.
Endotoxin Testing
We test water or samples of media to determine the level of endotoxin using the LAL (Lumilius Amebosite Lysate) gel clot assay from Associates of Cape Cod, MA. At least 200 µl samples should be submitted in endotoxin free container. Generally, radiation sterilized tissue culture plasticware is endotoxin free, but not autoclaved glassware. Three dilutions per sample are tested routinely, along with a set of blanks and positive controls.
EBV Transformation
Epstein Barr Virus (EBV) transformation is a reliable method to immortalize mammalian cells. It is most often used to obtain cell lines from human lymphocytes that serve as a permanent source for DNA isolation and protein isolation. The technique has found widespread use in clinical trials as the principal method of generating a permanent source of patient DNA for genotyping. Patient genotyping is becoming more popular as the genetic causes of diseases are being unraveled at increasing speed.
EBV is the causative agent of human infectious mononucleosis , a childhood disease. More than 90% of adults are a carrier of EBV. Adults usually show no EBV symptoms.
EBV is a Herpesvirus and its genome consists of a 172kb linear double stranded DNA which has been completely sequenced. EBV molecular biology and pathogenesis are extensively studied and the roles of many crucial EBV and host cell genes in pathogenesis are known. EBV infects only certain mammalian epithelial cells and B lymphocytes. In vitro EBV immortalizes B-cells by activating a number of cell cycle regulating genes as well as B-cell specific genes including immunoglobulin genes. Usually B-cell infections are latent and only strong stimulation can cause the lytic cycle. EBV DNA is replicated as an episomal ring and 10-20 copies per cell are typical.
IMPORTANT: When working with EBV (or any other infectious agent) negative control blood samples may NOT be obtained from laboratory personnel in order to minimize the risk of infections with one’s own EBV-transformed blood cells against which the body would not be able to mount an immune response.
Fly Food
The facility prepares media for Drosophila culture in vials and bottles using the method of Spradling. We also make a modified version where the molasses is replaced by dextrose.
Please contact Sabine Baxter for details.
