OBJECTIVE The validity of peak expiratory flow variation (PEFvar) as defined by National Heart, Lung, and Blood Institute (NHLBI) guidelines as a diagnostic tool for suspected asthma or its comparative value to methacholine inhalation challenge (MIC) or postbronchodilator (BD) FEV(1) responses has not been formally assessed. We prospectively analyzed the correlation of 28 different PEFvar indexes (including 4 NHLBI-compatible indexes) with MIC and pre-BD and post-BD FEV(1) responses in suspected asthmatic subjects with normal findings on lung examination, chest radiography, and baseline spirometry. METHODS Participants were asked to record peak expiratory flow four times daily for 2 to 3 weeks, followed by an MIC. During a minimum 6-month follow-up period, a clinical diagnosis of asthma was made or ruled out based on testing results and response to antiasthma therapy. METHODS Medical school-affiliated subspecialty private practice of allergy, asthma, and immunology. METHODS One hundred twenty-one suspected asthmatic patients with normal findings on lung examination, chest radiography, and baseline spirometry. RESULTS Fifty-seven subjects completed both the peak flow diary and the MIC and were accepted for statistical analysis. There were no statistically significant correlations between any peak expiratory flow index and MIC. Among the three diagnostic tools evaluated, MIC had the highest sensitivity (85.71%). All the PEFvar indexes and post-BD responses had low sensitivity and high false-negative rates. CONCLUSIONS PEFvar and post-BD FEV(1) responses are poor substitutes for MIC in the assessment of patients with suspected asthma with normal findings on lung examination, chest radiography, and spirometry. Our findings warrant a reconsideration of the NHLBI guidelines recommendation of the utility of PEFvar as a diagnostic tool for asthma in clinical practice.