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Slit Type <Cardiomyocyte>

Cell Culture Plate

Product code : CG 85 25 20

85μm(W)x25μm(H) 20μm(slit)

Cells

Cells were purchased from Primary Cell Co.,Ltd (Japan). These cells were isolated from 1-4 day old Sprague-Dawley (SD) rats.

Culture Medium

Culture media were purchased from Primary Cell Co.,Ltd (Japan). The medium contained inactivated fetal bovine serum (final concentration: 10%), penicillin G (10 IU/mL) and streptomycin (10 μg/mL) in Dulbecco's Modified Eagle Medium: Nutrient Mixture F-12 (DMEM/F-12).

Preparation of Coating Solution

0.02mol/L acetic acid was filtrated with 0.2μm pore-size membrane. The filtrated acetic acid and 4mg/mL collagen solution (BD Biosciences, Cat.Num.354236) were mixed as bellow.

0.02mol/L acetic acid (sterilized) : 9.937mL
4mg/mL collagen solution : 0.063mL
Total volume : 10mL

Coating

  1. 100μL of PBS was put in each well.
  2. PBS was aspirated.
  3. Before these wells were not dried, 65μL of 25μg/mL collagen solution were added in each well (5μg/cm2).
  4. The plates were incubated for an hour at room temperature.
  5. Solution was aspirated.
  6. The plates were washed twice by PBS. Coated plates were stored at 4℃ and used within a week.

Cell Seeding

  1. PBS was aspirated from each well of plates
  2. 80μL/well of cell suspension was added to Micro-Space Cell Culture Plates. (Cell density: 2x105 viable cells/mL (0.5x105 viable cells/cm2))
  3. Plates were gently shaken in a back-and-forth and side-to-side manner to disperse cells evenly in each well. Avoid cells’ clumping in the center of wells.
  4. Plates were carefully placed into a water-jacketed incubator at 37℃, 5% CO2, for one day.
  5. Culture medium was aspirated. Then, 80μL of pre-warmed (at 37℃) culture medium was added immediately to each well.
  6. Plates were placed carefully into a water-jacketed incubator at 37℃, 5% CO2.
  7. After cells were attached, culture medium was replaced by fresh culture medium every other day. Cells became confluent in a week.