Technical Specification: Cell Culture Reagents
1. Product Range
Solarbio offers a comprehensive range of cell culture reagents optimized for mammalian cell culture workflows, including phosphate-buffered saline formulations, enzymatic dissociation reagents, and culture media components. All products undergo stringent quality control for endotoxin, sterility, mycoplasma, and nuclease contamination.
| Product |
SKU |
Formulation |
Sterility |
Volume Options |
| D-PBS (Calcium and Magnesium Free) |
D1040 |
Dulbecco's PBS modified |
0.1 μm filtered |
500 mL, 1 L |
| D-PBS (with Ca²⁺/Mg²⁺) |
D1041 |
Dulbecco's PBS, standard |
0.1 μm filtered |
500 mL, 1 L |
| 10× D-PBS |
D1042 |
10× concentrate |
0.1 μm filtered |
500 mL, 1 L |
| Trypsin-EDTA (0.25%) |
T1300 |
Trypsin 2.5 g/L, EDTA 0.2 g/L |
0.1 μm filtered |
100 mL, 500 mL |
| Trypsin-EDTA (0.05%) |
T1305 |
Trypsin 0.5 g/L, EDTA 0.2 g/L |
0.1 μm filtered |
100 mL, 500 mL |
| PBS (10×), pH 7.4 |
P1020 |
Standard PBS concentrate |
0.1 μm filtered |
500 mL |
Dulbecco's PBS (D-PBS) is a balanced salt solution formulated to maintain physiological pH and osmolality during cell washing and handling. The distinction between calcium/magnesium-containing and calcium/magnesium-free formulations is critical: Ca²⁺/Mg²⁺-free D-PBS is used for cell dissociation and washing before trypsinization, while D-PBS with Ca²⁺/Mg²⁺ is used for cell handling steps where junctional integrity must be preserved.
| Component |
D-PBS without Ca²⁺/Mg²⁺ |
D-PBS with Ca²⁺/Mg²⁺ |
| NaCl |
8.0 g/L |
8.0 g/L |
| KCl |
0.2 g/L |
0.2 g/L |
| Na₂HPO₄·12H₂O |
2.89 g/L |
2.89 g/L |
| KH₂PO₄ |
0.2 g/L |
0.2 g/L |
| CaCl₂ (anhydrous) |
— |
0.133 g/L |
| MgCl₂·6H₂O |
— |
0.1 g/L |
| pH |
7.2–7.4 |
7.2–7.4 |
| Osmolality |
280–310 mOsm/kg |
280–310 mOsm/kg |
Trypsin, a pancreatic serine protease (EC 3.4.21.4), cleaves peptide bonds at the carboxyl side of lysine and arginine residues. In cell culture, trypsin hydrolyzes adhesion proteins (integrins, cadherins, fibronectin) that mediate cell-substrate and cell-cell attachment. EDTA chelates divalent cations (Ca²⁺, Mg²⁺) that are required for cadherin-dependent cell-cell adhesion and integrin-mediated cell-matrix adhesion, thereby enhancing trypsin dissociation efficiency.
The proteolytic reaction catalyzed by trypsin:
Trypsin
↓
Protein (adhesion molecules) → Peptide fragments + Free amino acids
(Lys/Arg - X bond hydrolysis)
| Parameter |
Trypsin-EDTA (0.25%) |
Trypsin-EDTA (0.05%) |
| Trypsin activity |
~2500 BAEE units/mL |
~500 BAEE units/mL |
| EDTA concentration |
0.02% (0.53 mM) |
0.02% (0.53 mM) |
| Recommended cell types |
HEK293, HeLa, A549, HepG2, Vero |
Primary cells, stem cells, sensitive lines |
| Incubation time (37°C) |
1–3 min |
3–5 min |
| Inactivation method |
Medium with serum or trypsin inhibitor |
Medium with serum or trypsin inhibitor |
| Neutralization pH |
7.2–7.4 |
7.2–7.4 |
| Storage |
-20°C, 18 months |
-20°C, 18 months |
4. Quality Specifications
| Parameter |
Specification |
Test Method |
| Endotoxin |
<0.5 EU/mL |
LAL kinetic assay |
| Sterility |
Sterile (0.1 μm filtration) |
Membrane filtration per USP <71> |
| Mycoplasma |
Not detectable |
qPCR and culture methods |
| pH |
7.2–7.4 |
Potentiometric |
| Osmolality |
280–310 mOsm/kg |
Freezing point depression |
| DNase/RNase |
Not detectable |
Fluorogenic substrate assay |
| Protease (non-trypsin products) |
Not detectable |
Azocasein assay |
| Heavy metals |
<10 ppm |
ICP-MS |
| Storage (D-PBS) |
2–30°C, 24 months |
— |
| Storage (Trypsin-EDTA) |
-20°C, 18 months |
— |
5. Recommended Protocols
Cell Washing (D-PBS)
| Step |
Detail |
| Remove culture medium |
Aspirate completely from monolayer |
| Add D-PBS |
1–2 mL per T25 flask or 200 μL per 96-well |
| Gently rinse |
Tilt flask/plate; avoid direct pipetting onto cell layer |
| Remove and repeat |
Aspirate; repeat for serum-containing cultures |
| Proceed |
Add trypsin or fresh medium as required |
Cell Dissociation (Trypsin-EDTA)
| Step |
Detail |
| Wash cells |
1× with Ca²⁺/Mg²⁺-free D-PBS to remove serum proteases |
| Add trypsin |
0.5 mL per T25 flask (0.25%); spread evenly |
| Incubate |
37°C for 1–3 min; monitor microscopically for rounding |
| Tap flask |
Gentle tapping to dislodge cells |
| Neutralize |
Add complete medium (2× trypsin volume) with 10% FBS |
| Centrifuge |
200–300 × g for 5 min; resuspend in fresh medium |
6. Cell Culture Applications and Reagent Selection
| Application |
Recommended Reagent |
Rationale |
| Routine cell passaging (HEK293, HeLa) |
D-PBS without Ca²⁺/Mg²⁺ (D1040) + Trypsin-EDTA 0.25% (T1300) |
Standard dissociation for robust adherent lines |
| Primary cell isolation |
D-PBS without Ca²⁺/Mg²⁺ (D1040) + Trypsin-EDTA 0.05% (T1305) |
Gentle dissociation preserves primary cell viability |
| Immunofluorescence staining |
D-PBS with Ca²⁺/Mg²⁺ (D1041) |
Maintains cell morphology during staining |
| Flow cytometry preparation |
D-PBS without Ca²⁺/Mg²⁺ (D1040) |
Prevents clumping; compatible with staining buffers |
| Cell washing before lysis |
D-PBS without Ca²⁺/Mg²⁺ (D1040) |
Avoids interference from Ca²⁺/Mg²⁺ in downstream assays |
| Intracellular staining |
D-PBS without Ca²⁺/Mg²⁺ (D1040) + fixation/permeabilization buffer |
Compatible with fixation protocols |
7. Cross-References
For product procurement: solarbio.store