BM Lin− (TER119, CD11b, Gr-1, CD3ε, NK1.1 or CD49b, Ly6C, Ly6G, CD11c)− CD19+ B cells were isolated from C57BL/CJ or BALB/CJ mice using FACSAriaTM Fusion sorter and 106/ml B cells were cultured in 50% cancer CM in cRPMI for 7 days in Nunc™ Multidishes with UpCell™ Surface (Thermo Fisher) without changing media for 7 days. 70z/3 pre-B cells (105/ml) were cultured in 50% cancer CM for up to 30 days with a replenishing culture medium every 3–4 days. Adherent cells (macrophages) were harvested by detaching them at 4 °C for 15 min in PBS. For Giemsa staining, B-MF was fixed with ethanol for 5 min, and Wright-Giemsa stained according to the manufacturer’s instructions. CSF1R receptor signaling was blocked with Ki20227 (R&D).
Nunc multidishes
Manufactured by Thermo Fisher Scientific
Sourced in United States
Nunc™ Multidishes are a line of sterile cell culture vessels designed for a variety of cell-based applications. They feature multiple individual culture wells on a single plate, allowing for parallel experiments or multiple cell lines to be cultured simultaneously. The products are available in different well sizes and formats to accommodate different experimental needs.
Automatically generated - may contain errors
3 protocols using nunc multidishes
1
Isolation and Culture of Tumor-Educated B Cells
2
Rapamycin Cytotoxicity in HUVEC-Pericyte Co-Culture
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HUVEC and pericytes were brought to confluence on 12 well Nunc multidishes (Thermo Scientific, Waltham, MA, USA), kept in growth-factor reduced medium for 24h to stop proliferation and treated with vehicle (DMSO; control) or increasing concentrations of rapamycin at 0.005, 0.05, 0.5, 1, 2.5, 5, 7.5, 10, 12.5 and 15 μg/ml, respectively. After 72 hours of treatment, the XTT-Assay was performed as described below. Differences in absorption compared to the untreated control were interpreted as differences in metabolic activity and cell count due to potential toxicity of rapamycin. All experiments were conducted on passages 3-7, performed as duplicates and repeated at least three times on different days.
3
Porcine Oocyte Maturation Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The maturation medium was porcine oocyte medium (POM; Research Institute for the Functional Peptides, Yamagata, Japan) supplemented with 1 mM dibutyryl cAMP (dbcAMP; Sigma-Aldrich), 10
ng/ml epidermal growth factor (EGF, Sigma-Aldrich), 10 IU/ml eCG (Serotropin; ASKA Pharmaceutical, Tokyo, Japan) and 10 IU/ml hCG (500 units; Puberogen, Novartis Animal Health, Tokyo,
Japan). Forty to fifty COCs were cultured in each well of 4-well dishes (Nunc MultiDishes, Thermo Fisher Scientific, Waltham, MA, USA) in 500 -µl droplets of IVM medium without oil coverage
for 22 h in an atmosphere of 5% CO2, 5% O2, and 90% N2 at 38.5°C. Thereafter, COCs were cultured in maturation medium without dbcAMP for an additional 22−24
h under the same atmosphere. At the end of IVM, COCs were denuded by treatment with 0.1% (w/v) hyaluronidase in collection medium for 2 min, followed by gentle pipetting through a
narrow-bore glass capillary tube in collection medium without hyaluronidase. Thereafter, oocytes were investigated under a stereo microscope. Those with an intact oolemma, a normal spherical
shape, a smooth surface, and a dark and evenly granular cytoplasm were considered to be alive. Oocytes with membrane damage and a brownish faded cytoplasm were considered dead. Survival
rates are expressed as the living oocytes on the total number of oocytes.
ng/ml epidermal growth factor (EGF, Sigma-Aldrich), 10 IU/ml eCG (Serotropin; ASKA Pharmaceutical, Tokyo, Japan) and 10 IU/ml hCG (500 units; Puberogen, Novartis Animal Health, Tokyo,
Japan). Forty to fifty COCs were cultured in each well of 4-well dishes (Nunc MultiDishes, Thermo Fisher Scientific, Waltham, MA, USA) in 500 -µl droplets of IVM medium without oil coverage
for 22 h in an atmosphere of 5% CO2, 5% O2, and 90% N2 at 38.5°C. Thereafter, COCs were cultured in maturation medium without dbcAMP for an additional 22−24
h under the same atmosphere. At the end of IVM, COCs were denuded by treatment with 0.1% (w/v) hyaluronidase in collection medium for 2 min, followed by gentle pipetting through a
narrow-bore glass capillary tube in collection medium without hyaluronidase. Thereafter, oocytes were investigated under a stereo microscope. Those with an intact oolemma, a normal spherical
shape, a smooth surface, and a dark and evenly granular cytoplasm were considered to be alive. Oocytes with membrane damage and a brownish faded cytoplasm were considered dead. Survival
rates are expressed as the living oocytes on the total number of oocytes.
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!