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We performed in-situ measurements of mechanical properties of individual W303 wild-type yeast cells by using a soft nanoneedle inside an integrated environmental scanning electron microscope (ESEM) - nanomanipulator system. Two different spring constants of soft nanoneedles which can buckle from certain applied compression force were used to measure a general local stiffness, i.e. stiffness property of the whole cell from a single indentation point, and a finer local stiffness, i.e. stiffness property of a cell surface from a single indentation point. The finer local stiffness property of the single cells was also being compared at different pressure and different temperature ranges. The advantages of using a soft nanoneedle for single cells stiffness characterizations are severe cell damage from an excessive indentation force could be prevented and big variations in single cell stiffness property could be easily detected by examining either from the dented mark area of the local indentation point or by comparing the buckling length of the nanoneedle. The soft nanoneedle which currently used for measuring single cells stiffness variations could be applied in the future fast disease diagnosis based on single cell stiffness analysis.