Stable isotopes in tree-ring α-cellulose are valued as environmental proxies and their use is steadily increasing; however, preparation of α-cellulose is a bottleneck in isotope analysis. Recent methodological breakthrough for extracting tree-ring α-cellulose directly from tree-ring cross-sectional laths drastically increased the throughput of tree-ring isotope data. In this paper, we evaluate our recently designed “cross-section” method. This method employs polytetrafluoroethylene (PTFE) cases, enabling direct extraction of α-cellulose from 1-mm thick tree-ring laths, in combination with fixation sheets to prevent disintegration of freeze-dried α-cellulose laths. Perforated PTFE cases are easily producible at an affordable cost. They are made of commonly available lab consumables in catalogs and do not require specially made PTFE parts. Freeze-dried α-cellulose laths preserved distinct anatomical structure, enabling precise separation at the tree-ring boundaries. Once separated from a lath, tree-ring α-cellulose can be weighed directly into silver or tin capsules for analysis. We checked chemical purity of α-cellulose prepared by the cross-section method from five tree species (larch, pine, spruce, beech, and oak). Residual lignin and hemicellulose contents were quantitatively assessed by Fourier transform infrared spectrometry and gas chromatography. The average chemical purity of α-cellulose laths from the five species was 94.5%, similar to the chemical purity of α-cellulose prepared with the standard Jayme-Wise method. Both oxygen and carbon isotope values of α-cellulose prepared by the cross-section method also closely matched those prepared by the standard method. We conclude that, by overhauling the method of α-cellulose preparation for tree-ring isotope analysis, we increased throughput of tree-ring oxygen and carbon isotope data without sacrificing sample purity. •We improved the complete process of α-cellulose extraction from tree rings for isotope analysis in order to enhance its throughput considerably.•Our PTFE case enabled extraction of α-cellulose from tree-ring plates as thin as 0.5–1.0mm.•Fixing freeze-dried cellulose plates onto sheets made handling of fragile cellulose plates easier.•The average purity of α-cellulose prepared with our method was 94.5%, which is similar to the standard method.