From: michalp717@gmail.com   
      
   Goal:    
   -Need to generate a pH vs. Fluorescent Intensity calibration curve using a   
   spectrofluorometer for a particular fluorescent dye with an analyte (metal   
   ion).   
      
   Method:   
   -Generating pairs of samples at varying pH (9, 9.5, 10, etc) with one sample   
   being a blank and the other containing the analyte.    
   1. First mix the dye with ethanol and HNO3 in a beaker to dissolve.    
   2. Next, 2.0 mL of this dye solution is added to two cuvettes as a pair. This   
   is repeated for varying pHs by adding base.    
   3. Finally, 0.1 mL of the analyte suspended in 3% or 0.3% nitric acid   
   (required to solubilize) is added to one cuvette (test sample), and 3% or 0.3%   
   nitric acid is added to the other cuvette (blank sample). This changes the pH   
   yet again, so it's measured    
   in the cuvette. Of course, the cuvette pH is always lower than that of the   
   beaker since the analyte is in acidic solution.   
      
   Problem:   
   -The analyte can only be managed in a separate facility, so I cannot add it to   
   the beaker solution. It must be added to the cuvette in the final step.    
   This creates the issue: I cannot adjust the pH in the cuvette given its small   
   volume, so whatever pH it ends up being after addition of the analyte, I have   
   to accept. This prevents me from adjusting the final pH to my desired outcome.    
   -This is made even more difficult with the equivalence point, preventing me   
   from reaching pHs of ~10 and 11 since the addition of the analyte in acid   
   results in a drastic pH decrease and even 0.1 mL is proportionally large   
   compared to 2.0 mL of the dye    
   solution..   
      
   Attempts to resolve:   
   -Tried using the analyte suspended in 0.3% acid solution rather than 3% acid   
   solution, but this doesn't fix the problem, only reduces the decrease in pH.    
      
   Ideally, whatever the pH of my beaker solution is, I could just add that to   
   the cuvettes that contains the analyte and the dye solution. Is this really   
   the only way? I feel I have been using my method for so long that I do not see   
   another way.   
      
   Thanks for reading   
      
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