Cellulose-based　separator　exhibits　excellent　electrolyte　affinity,　thermal　stability,　and　mechanical　strength,　which　acts　as　a　promising　alternative　to　commercial　polyolefin　separators　in　lithium　metal　batteries　(LMBs).　Fiber　size　in　cellulose-based　separators　plays　a　crucial　role　in　determining　their　physicochemical　structure　and　mechanical　strength,　as　well　as　the　electrochemical　performance　of　corresponding　LMBs.　Herein,　the　fiber　size　in　cellulose-based　separators　was　first　time　regulated　to　optimize　their　mechanical　stability　and　the　related　battery　performance.　The　influences　of　fiber　size　in　the　separator　on　chemical　structure,　mechanical　properties,　surface　morphology,　electrochemical　behavior　were　investigated　in　detail,　in　which　the　underlying　mechanism　between　separator　structure　and　the　related　performance　was　elucidated.　As　a　result,　the　separator　optimized　by　fiber　size　regulation　exhibited　excellent　thermal　stability　under　180　°C,　good　tensile　strengths　of　6.0　MPa　and　Young＇s　moduli　of　315.9　MPa,　superior　room　temperature　ionic　conductivity　of　1.87　mS　cm−1,　as　well　as　significantly　improved　electrochemical　performance　of　corresponding　batteries.　It　can　be　concluded　that　structure　optimization　for　cellulose-based　separator　through　fiber　size　regulation　is　an　effective　and　indispensable　approach　towards　high　safety　and　high　performance　LMBs.　©　2024　Wiley-VCH　GmbH.