The development of scaffold materials that effectively mimic the extracellular matrix while enabling controlled nutrient delivery remains a critical challenge in tissue engineering. Multi-channel collagen gels (MCCGs), which form through the competition between gelation and phase separation, have emerged as promising scaffolds due to their self-organized vessel-like structures. However, a systematic understanding of the relationship between the gelation conditions and functional properties is limited. In this study, MCCGs were developed as controllable perfusion culture scaffolds by investigating the effects of carbonate buffer concentration on channel formation, permeation behavior, and cell proliferation. MCCGs were prepared using different carbonate buffer concentrations (12.5, 25, and 50 mM), with 25 mM producing optimal channel formation, characterized by an approximately 60% channel area fraction and uniform distribution. Permeation studies revealed that fluid transport through MCCGs is governed by a complex interplay between capillary phenomena and hydraulic pressure, whose relative dominance shifts with flow rate: capillary action dominates at low flow rates (2.5 mL/h), whereas hydraulic pressure becomes the primary driver at higher rates (5.0-10.0 mL/h). Cell proliferation assessments demonstrated that MCCGs prepared with 25 mM carbonate buffer provided the most favorable microenvironment, achieving superior cell growth over 168 h through balanced media supply and cell adhesion area. This optimization approach through buffer concentration adjustment offers a cost-effective and scalable method for developing perfusion culture scaffolds, advancing both the fundamental understanding of functional gel systems and practical applications in tissue engineering and regenerative medicine.