A method for validating a PCR instrument, the method comprising: receiving amplification data from a validation plate to generate a plurality of amplification curves, wherein a first portion of wells of the validation plate includes a sample of a first quantity and another portion of the wells of the validation plate includes a second quantity of the sample, wherein the first quantity and the second quantity are different, and each amplification curve includes an exponential region; determining a set of fluorescence thresholds by evenly dividing by a predetermined number a range of fluorescence values within the exponential regions of the plurality of amplification curves, the dividing being performed to generate, for each of the amplification curves, a set of fluorescence values for the set of fluorescence thresholds, the range of fluorescence values being determined from a lowest fluorescent value of a bottom of the exponential regions of the plurality of amplification curves to a highest fluorescent value of a top of the exponential regions of the plurality of amplification curves; determining, for each fluorescence threshold of the set for each of the amplification curves, a first set of cycle threshold values C tquant1 of amplification curves generated from the samples of the first quantity and a second set of cycle threshold values C tquant2 of amplification curves generated from the samples of the second quantity; and calculating if the first and second quantities are sufficiently distinguishable based on the cycle threshold values at each of the plurality of fluorescence thresholds, wherein the calculating if the first and second quantities are sufficiently distinguishable includes determining a mean μ and a standard deviation σ of the first and second sets of cycle threshold values, wherein the first and second quantities are sufficiently distinguishable if the following equation results in a positive number for at least one fluorescence threshold: μC tquant 1 − 3 σC tquant 1 − μC tquant 2 + 3 σC tquant 2. receiving amplification data from a validation plate to generate a plurality of amplification curves, wherein a first portion of wells of the validation plate includes a sample of a first quantity and another portion of the wells of the validation plate includes a second quantity of the sample, wherein the first quantity and the second quantity are different, and each amplification curve includes an exponential region; determining a set of fluorescence thresholds by evenly dividing by a predetermined number a range of fluorescence values within the exponential regions of the plurality of amplification curves, the dividing being performed to generate, for each of the amplification curves, a set of fluorescence values for the set of fluorescence thresholds, the range of fluorescence values being determined from a lowest fluorescent value of a bottom of the exponential regions of the plurality of amplification curves to a highest fluorescent value of a top of the exponential regions of the plurality of amplification curves; determining, for each fluorescence threshold of the set for each of the amplification curves, a first set of cycle threshold values C tquant1 of amplification curves generated from the samples of the first quantity and a second set of cycle threshold values C tquant2 of amplification curves generated from the samples of the second quantity; and calculating if the first and second quantities are sufficiently distinguishable based on the cycle threshold values at each of the plurality of fluorescence thresholds, wherein the calculating if the first and second quantities are sufficiently distinguishable includes determining a mean μ and a standard deviation σ of the first and second sets of cycle threshold values, wherein the first and second quantities are sufficiently distinguishable if the following equation results in a positive number for at least one fluorescence threshold: μC tquant 1 − 3 σC tquant 1 − μC tquant 2 + 3 σC tquant 2.