To construct an anti-αvβ8 antibody with reduced Fc receptor binding and effector function the variable regions from ADWA11 were subcloned into a mIgG1 Fc backbone that contained E233P, E318A, K320A, and R322A mutations, as outlined in https://patents.google.com/patent/US20090155256 . ADWA-11_4mut binding to mouse αvβ8 was assessed using C8-D1A mouse astrocyte cells (ATCC, CRL2541) and blockade of TGFβ activation was determined using C8-S cells in a co-culture TMLC luciferase assay, described above.
Crl 2541
Manufactured by American Type Culture Collection
Sourced in United States
The CRL-2541 is a type of laboratory equipment. It is a generic model number and does not provide detailed information about the specific function or capabilities of the equipment.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin, by Thermo Fisher Scientific (4 mentions)
Streptomycin, by Thermo Fisher Scientific (4 mentions)
Crl 2299, by American Type Culture Collection (3 mentions)
L glutamine, by Thermo Fisher Scientific (2 mentions)
C57bl 6, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Crl 2540, by American Type Culture Collection (1 mentions)
Humidified incubator, by NuAire (1 mentions)
Hb 8065, by American Type Culture Collection (1 mentions)
Amphotericin b, by Thermo Fisher Scientific (1 mentions)
Ccl 81, by American Type Culture Collection (1 mentions)
Sodium pyruvate, by Thermo Fisher Scientific (1 mentions)
9 protocols using crl 2541
1
Anti-αvβ8 Antibody Engineering
2
BBB model from bEnd.3 and C8D1A cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BBB model was obtained by culturing the brain-derived endothelioma bEnd.3 cells (ATCC® CRL-2299™ at 8 × 104 cells/cm2) and the C8D1A brain astrocytes (ATCC® CRL-2541™ at 2 × 104 cells/cm2) on the luminal and abluminal side of 3 μm porous transwells (Corning Incorporated). Expression of tight junction proteins and BBB crossing were assessed as described in Supplementary methods .
3
In Vitro BBB Model with Endothelial, Astrocyte and Pericyte
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For mouse brain endothelial cells (bEnd.3, ATCC® CRL-2299™), the medium used was DMEM supplemented with 10% FCS, penicillin and streptomycin, L-glutamine and Fungizone. Astrocyte (ATCC® CRL-2541™, C8-D1A Astrocyte Type I clone) medium was antibiotics free DMEM supplemented with 10% FCS and L-glutamine. Pericyte (MSC, Gibco®iMouse, C57BL⁄6) medium used was DMEM F12 media with gluta-MAX-I, supplemented with 10% FCS and 5 μg/ml gentamicin. For transwell experiments, both sides of the transwell insert filters (Corning®3460 PE filter, diameter: 1.05 cm, pore size: 0.4μm) were pre-coated with 10 μg/cm2 collagen for 2 hours at room temperature. This was followed by seeding bEnd.3 endothelial cells on the upper surface of the transwell at a density of 20,000–40,000 cells/well, and incubated for 12 hours at 37 °C in 95% air 5% CO2 in order to allow the cells to fully attach. Next, the astrocytes and/or pericytes (10,000–20,000 cells/well) were seeded on the opposite side of the filter insert, and incubated for 12 hours at 37 °C in 95% air 5% CO2. Finally the inserts were moved to a transwell plate, and incubated for 7 days at 37 °C, the medium being changed every two days.
4
Anti-αvβ8 Antibody Engineering
5
In Vitro Cortical Spreading Depolarization
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Murine brain endothelial cells, bEnd.3 (CRL-2299, ATCC) were cultured in DMEM (Gibco, Sigma Aldrich, St Louis, MO, USA 11995-065), supplemented with 2 mM L-glutamine (ThermoScientific, 25030081), 10% fetal bovine serum (Gibco, 10082139), and penicillin (100 UI/mL)-streptomycin (100 μg/mL) (Invitrogen, 15140122). Cells were cultured at 37 °C in a humidified 5% CO2/95% air atmosphere and were grown to 80% confluence in all experiments. Twenty-four hours before treatment, the media of bEnd.3 cells were changed to astrocyte-conditioned media (ACM) harvested from confluent C8-D1A flasks. The C8-D1A cells (CRL-2541, ATCC) were cultured in DMEM (Gibco, 11995-065), supplemented with 10% fetal bovine serum and penicillin-streptomycin. To mimic CSD event in vitro, bEnd.3 cells were treated with 100 mM KCl for 5 min, which is a typical condition to evoke potassium-triggered spreading depolarization in live brain slices [44 (link)].
6
Cultivation and Characterization of Murine Microglia and Astrocytes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Murine C8-B4 microglia cells (ATCC CRL-2540; monocytes from cerebellum of C57BL/6 mice (Mus musculus) and C8-D1A astrocytes (ATCC CRL-2541; astrocyte type I clone from C57BL/6 mouse cerebellum) were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA) and grown in DMEM-supplemented with 10% FBS and 1% penicillin/streptomycin. The microglia and astrocytes were cultured at 37 °C under 5% CO2 in a humidified incubator (NuAire, Plymouth, MN, USA) and sub-cultivated by trypsination once/twice per week in a 1:5/1:10 split ratio, respectively. They were allowed to grow as monolayers in 75 cm2 cell culture flasks with filter screwcaps (Techno Plastic Products, Trasadingen, Switzerland) until 80–100% confluence was reached. The microglia and astrocytes were plated at 90,000/cm2 and 45,000/cm2, respectively, 24 h prior to the test, which resulted in approximately 90% confluence at the day of exposure as observed by light microscopy (Leica DMIL, Solms, Germany). The cell culture medium was refreshed before performing the exposure test.
7
Culturing Diverse Cell Lines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Astrocyte cell line (ATCC, CRL-2541), human glioblastoma astrocytoma cell line U373 (ATCC, HTB17-1055), and human hepatoma HepG2 cells (ATCC, HB-8065) were used in the study. The cells were cultured in standard conditions (37 °C, 5% CO2), in EMEM medium with 2 mM glutamine and 1% non-essential amino acids, all supplemented with 10% FBS and antibiotics (penicillin–streptomycin; 10,000 U/mL; final concentration in culture medium was 1%).
8
Mouse Astrocyte Culture for TBI Studies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The mouse astrocytes (CRL‐2541) used in this project were purchased from American Type Culture Collection (ATCC, Manassas, VA) because of their known utility in TBI studies.[45 ] These cells are specifically clonal permanent cell lines with astrocytic properties, instead of being solely an astrocyte culture. This culture provides a solid foundation for the response that neuronal cells have to cavitation and future studies aim to use primary astrocyte cultures. The cells were grown according to company protocol in growth media (ATCC, 30‐2002). The culture media was also supplemented with 10% FBS and 1% penicillin (10 000 U mL−1)‐ streptomycin (10 000 µg mL−1), both of which were purchased from Gibco (Waltham, MA). The cells in all of the data collection consist of culture passage numbers 3–7. The astrocytes were housed in an incubator set at 37 °C with 5% CO2.
9
Murine Astrocyte and Vero Cell Culture
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Murine astrocyte C8-D1A and African green monkey kidney (Vero) cell lines were purchased from the American Type Culture Collection (ATCC® CRL-2541™ and ATCC® CCL-81™, respectively). C8-D1A cells were grown in Dulbecco's modified essential medium (D-MEM), supplemented with 10% fetal bovine serum (FBS), 4 mM L-glutamine, 1 mM sodium pyruvate (Gibco by Thermo Fisher Scientific, Carlsbad, CA, USA), 5 g/l glucose, 100 U/ml penicillin, 100 μg/ml streptomycin, and 0.25 μg/ml amphotericin B (Thermo Fisher Scientific) in standard conditions. Vero cells were grown in Eagle's minimum essential medium, supplemented with 10% FBS, 100 U/ml penicillin, 100 μg/ml streptomycin, and 0.25 μg/ml amphotericin B (Thermo Fisher Scientific).
HSV-1 strain McKrae [24 (link)] was grown (PFU/ml) in African green monkey kidney (Vero) cells. Virus titers were determined by plaque assay on Vero cells.
HSV-1 strain McKrae [24 (link)] was grown (PFU/ml) in African green monkey kidney (Vero) cells. Virus titers were determined by plaque assay on Vero cells.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!