As an extension of recent results (Rhodes, Xu and Bittman (1992) Biochim. Biophys. Acta 1128, 93; Hui, Xu and Bittman (1992) Langmuir 8, 2724) with a C18 diacetylenic phosphatidylcholine, bilayers of 1,2-bis(pentacosa-4,6-diynoyl)-sn-glycero-3-phosphocholine (C25) were investigated using X-ray diffraction on multibilayers and electron diffraction on Langmuir-Blodgett deposited bilayers. Monolayers of this lipid form solid (gel phase) domains at pi > 14 mN/m. Electron diffraction data indicated that the chain spacing in these domains is 4.25 A and that the chains are tilted at angle of approximately 35 degrees relative to the bilayer plane. Wide angle data from X-ray diffraction experiments indicated a similar spacing and chain tilt. Small angle data showed that the lamellar repeat was 70 A at high humidity and < or = 60 A at low humidity. The bilayer electron density profiles indicated a bilayer structure with no interdigitation. High angle reflections indicate that the principal acyl chain repeat is preserved as a function of hydration but some rearrangement occurs for other reflections. The approximately 10 A reflection corresponding to the headgroup spacing previously observed with C18-diacetylenic phosphatidylcholine bilayers was not observed. The results are interpreted in terms of a packing model and possible limitations or constraints to the polymerization process.