Lanolin, also known as wool wax, is derived from sheep and has diverse applications in food, cosmetic, textile and lubricant industries. Owing to its direct contact with human, there is a need of studying pesticide residues as contaminants in lanolin. The present study describes a single novel hyphenated gas chromatography-tandem mass spectrometry (GC-MS/MS) technique for the quantification of 14 organophosphorus (OP) pesticides (tecnazene, propetamphos, diazinon, dichlofenthion, chlorpyrifos methyl, fenchlorphos, malathion, chlorpyrifos, pirimiphos ethyl, bromophos ethyl, tetrachlorvinphos, ethion, phosalone, and coumaphos) in lanolin using electrospray ionization (EI) with multiple reaction monitoring (MRM) acquisition mode. The method is simple in terms of sample preparation steps based on matrix solid-phase dispersion (MSPD). The method was found to be linear over the analytical range of 0.05-2.0 μg/g, with acceptable coefficient of determination (r 2 ≥ 0.99) for all the 14 pesticides. The limit of detection (LOD) and limit of quantification (LOQ) of the method were found to be less than 0.05 and 0.1 μg/g, respectively, for all the 14 OP pesticide residues. The precision and accuracy of the method were found to be within the acceptable limits, that is, recoveries in the range of 83.5%-104.1% with less than 12.5% of relative standard deviation for all the pesticides. The multiresidue method for estimating pesticide residues employing GC-MS/MS technique will be useful for OP pesticide levels in large number of lanolin samples.