{"text_format":"2","workbook":"xlsx","n_sheets":3,"sheets":["Experiment_example","Results_example","Calculation-example"],"not_rendered":{"charts":0,"images":0,"macros":0,"external_links":0,"defined_names":0,"comments":0,"pivot_tables":0},"unopened":[],"bounds":{"document_bytes":16777216,"sheet_cells":100000,"run_cost":"one","rows_first":2000,"rows_last":500,"rows_between":2000,"halvings":0,"cell_chars":16384,"cell_head_chars":4096},"cells_cut":0,"cells_raw":0,"read":"whole","date_system":"1900"}

{"sheet":"Experiment_example","index":0,"n_rows":37,"n_cols":13,"rows_kept":37,"cut":false,"hidden":false,"merged":[]}
[null,null,"Stock concentration (in μM)","Final concentration  (in μM)","Total voume "]
["Enviromental Mastermix",null,2,1,{"$f":"(B12+B10)*B11*D2/C2","v":900}]
["Probe CMV",null,100,0.3,{"$f":"(B12+B10)*B11*D3/C3","v":5.4}]
["Primer CMV+",null,100,1,{"$f":"(B12+B10)*B11*D4/C4","v":18}]
["Primer CMV-",null,100,1,{"$f":"(B12+B10)*B11*D5/C5","v":18}]
[]
[]
["Water",null,null,null,{"$f":"(B12*B11)-(SUM(E2:E5))","v":498.6}]
["vol mix 1x",null,"total",{"$f":"E11+B10","v":10},{"$f":"SUM(E2:E8)","v":1440}]
["Volume of template per well",10]
["Number of wells*",36]
["Volume of reagent per well",40]
["* always add 10% additional wells to account for loss of reagents during pipetting"]
[]
[]
[]
[null,1,2,3,4,5,6,7,8,9,10,11,12]
["A","D5","D5","Sample 1","Sample 1"]
["B","D6","D6","Sample 2","Sample 2"]
["C","D7","D7","Sample 3","Sample 3"]
["D","D8","D8","Sample 4","Sample 4"]
["E","D9","D9","Sample 5","Sample 5"]
["F","D10","D10","Sample 6","Sample 6"]
["G","D11","D11","Sample 7","Sample 7"]
["H","D12","D12","H2O","H2O"]
[]
[]
[]
["Program:",null,null,null,"STANDARD ","Dil factor","Template","Tris","Expected copy number/wells for CMV"]
[null,"50˚C","2 mins",null,"D5",null,null,null,800000]
[null,"95˚C","10 mins",null,"D6",10,5,45,{"$f":"I30/10","v":80000}]
["Then run 50 cycles at:",null,null,null,"D7",10,5,45,8000]
[null,"95˚C","15 secs",null,"D8",10,5,45,800]
[null,"60˚C","1 mins",null,"D9",10,5,45,80]
[{"$nulls":4},"D10",10,5,45,8]
[{"$nulls":4},"D11",10,5,45,0.8]
[{"$nulls":4},"D12",10,5,45,0.08]

{"sheet":"Results_example","index":1,"n_rows":24,"n_cols":10,"rows_kept":24,"cut":false,"hidden":false,"merged":[[0,1,0,3],[0,4,1,4],[0,7,1,7],[0,8,1,8],[0,9,1,9]]}
[null,"Standard",null,null,"Copies/well",null,null,"Sample ID","Wells","Ct for sample"]
[null,"Dilution","Well","CT"]
[null,"D5","A1",20.580317000000001,800000,null,null,"Sample  1","A3",30.422132000000001]
[null,null,"A2",20.413937000000001,{"$nulls":4},"A4",30.165602]
[null,"D6","B1",23.446764000000002,80000,null,null,"Sample  2","B3",32.388460000000002]
[null,null,"B2",23.264230000000001,{"$nulls":4},"B4",32.662604999999999]
[null,"D7","C1",25.867923999999999,8000,null,null,"Sample 3","C3","Undetermined"]
[null,null,"C2",25.940131999999998,{"$nulls":4},"C4","Undetermined"]
[null,"D8","D1",29.378288000000001,800,null,null,"Sample 4","D3",35.691135000000003]
[null,null,"D2",29.40943,{"$nulls":4},"D4",35.799999999999997]
[null,"D9","E1",32.816764999999997,80,null,null,"Sample 5","E3","Undetermined"]
[null,null,"E2",32.826590000000003,{"$nulls":4},"E4","Undetermined"]
[null,"D10","F1",36.527985000000001,8,null,null,"Sample 6","F3","Undetermined"]
[null,null,"F2",35.376919999999998,{"$nulls":4},"F4","Undetermined"]
[null,"D11","G1",48.611904000000003,0.8,null,null,"Samples 7","G3",35.022796999999997]
[null,null,"G2",37.809049999999999,{"$nulls":4},"G4",35.599580000000003]
[null,"D12","H1","Undetermined",0.08,null,null,"H2O","H3","Undetermined"]
[null,null,"H2","Undetermined",{"$nulls":4},"H4","Undetermined"]
[]
[]
[]
[null,"NOTE:","Consider repeating samples with differences of 0.3 Cts between duplicates. Run triplicates whenever possible to improve precision. "]
[null,null,"Samples at the lower limit of detection might have discordant Ct values due to Poisson distribution of templates across wells"]
[null,null,"Thus, for a reliable low copy detection, a large number of replicates are necessary to provide statistical significance and to overcome the Poisson distribution limitation."]

{"sheet":"Calculation-example","index":2,"n_rows":37,"n_cols":15,"rows_kept":37,"cut":false,"hidden":false,"merged":[[15,11,16,11],[15,12,16,12],[15,13,16,13],[15,14,16,14]]}
["Sample ID","Replicate","µl-orig template","Dil-factor","Copy number/wells","log-lys","ct Values","Slope (a)","Intercept (b)","R2"]
["CMV Standard","D5",10,1,800000,{"$f":"LOG(E2)","v":5.9030899869919438},20.580317000000001,{"$f":"LINEST(G2:G13,F2:F13)","v":-3.1189999999999993},{"$f":"INTERCEPT(G2:G13,F2:F13)","v":38.601677836094538},{"$f":"RSQ(G2:G13,F2:F13)","v":0.99535294347004244}]
["CMV Standard","D5",10,1,800000,{"$f":"LOG(E3)","v":5.9030899869919438},20.413937000000001,{"$f":"H2","v":-3.1189999999999993},{"$f":"I2","v":38.601677836094538},{"$f":"$J$2","v":0.99535294347004244}]
["CMV Standard","D6",10,10,80000,4.9030899869919438,23.446764000000002,{"$f":"H3","v":-3.1189999999999993},{"$f":"I3","v":38.601677836094538},{"$f":"$J$2","v":0.99535294347004244},null,"y = ax + b"]
["CMV Standard","D6",10,10,80000,4.9030899869919438,23.264230000000001,-3.1189999999999993,38.601677836094538,0.99535294347004244,null,"a = slope obtained from the standard curve."]
["CMV Standard","D7",10,100,8000,3.9030899869919438,25.867923999999999,-3.1189999999999993,38.601677836094538,0.99535294347004244,null,"b = intercept obtained from the standard curve. "]
["CMV Standard","D7",10,100,8000,3.9030899869919438,25.940131999999998,-3.1189999999999993,38.601677836094538,0.99535294347004244,null,"y = Threshold Ct value for seminal sample."]
["CMV Standard","D8",10,1000,800,2.9030899869919438,29.378288000000001,-3.1189999999999993,38.601677836094538,0.99535294347004244,null,"x = Extrapolated amount of target DNA or cDNA for seminal sample."]
["CMV Standard","D8",10,1000,800,2.9030899869919438,29.40943,-3.1189999999999993,38.601677836094538,0.99535294347004244]
["CMV Standard","D9",10,10000,80,1.9030899869919435,32.816764999999997,-3.1189999999999993,38.601677836094538,0.99535294347004244]
["CMV Standard","D9",10,10000,80,1.9030899869919435,32.826590000000003,-3.1189999999999993,38.601677836094538,0.99535294347004244]
["CMV Standard","D10",10,100000,8,0.90308998699194354,36.527985000000001,-3.1189999999999993,38.601677836094538,0.99535294347004244]
["CMV Standard","D10",10,100000,8,0.90308998699194354,35.376919999999998,-3.1189999999999993,38.601677836094538,0.99535294347004244]
["CMV Standard","D11",10,1000000,0.8,{"$f":"LOG(E14)","v":-9.6910013008056392E-2},48.611904000000003,-3.1189999999999993,38.601677836094538,0.99535294347004244]
["CMV Standard","D11",10,1000000,0.8,{"$f":"LOG(E15)","v":-9.6910013008056392E-2},37.809049999999999,-3.1189999999999993,38.601677836094538,0.99535294347004244]
["CMV Standard","D12",10,10000000,0.08,{"$f":"LOG(E16)","v":-1.0969100130080565},"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,null,"Copies per well*","Copy per elution","Average of duplicate wells","Copies per ml seminal plasma***"]
["CMV Standard","D12",10,10000000,0.08,{"$f":"LOG(E17)","v":-1.0969100130080565},"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244]
["Sample 1",1,10,null,null,null,30.422132000000001,{"$f":"H17","v":-3.1189999999999993},{"$f":"I17","v":38.601677836094538},0.99535294347004244,{"$f":"(G18-I18)/H18","v":2.6224898480585246},{"$f":"POWER(10,K18)","v":419.26619592542232},{"$f":"L18*10","v":4192.6619592542229},{"$f":"AVERAGE(M18:M19)","v":4629.7519880730506},{"$f":"N18*5","v":23148.759940365253}]
["Sample 1",2,10,null,null,null,30.165602,-3.1189999999999993,38.601677836094538,0.99535294347004244,{"$f":"(G19-I19)/H19","v":2.7047373632877654},{"$f":"POWER(10,K19)","v":506.68420168918772},{"$f":"L19*10","v":5066.8420168918774}]
["Sample 2",1,10,null,null,null,32.388460000000002,{"$f":"H19","v":-3.1189999999999993},38.601677836094538,0.99535294347004244,1.9920544520982806,{"$f":"POWER(10,K20)","v":98.18710428731039},{"$f":"L20*10","v":981.87104287310387},{"$f":"AVERAGE(M20:M21)","v":891.92160247956224},{"$f":"N20*5","v":4459.6080123978109}]
["Sample 2",2,10,null,null,null,32.662604999999999,-3.1189999999999993,38.601677836094538,0.99535294347004244,1.9041592933935685,{"$f":"POWER(10,K21)","v":80.197216208602057},801.9721620860206]
["Sample 3",1,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0,{"$f":"AVERAGE(M22:M23)","v":0},{"$f":"N22*5","v":0}]
["Sample 3",2,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0]
["Sample 4",1,10,null,null,null,35.691135000000003,-3.1189999999999993,38.601677836094538,0.99535294347004244,0.93316538508962354,{"$f":"POWER(10,K24)","v":8.5736427874186045},85.736427874186049,{"$f":"AVERAGE(M24:M25)","v":82.42596326767702},{"$f":"N24*5","v":412.12981633838513}]
["Sample 4",2,10,null,null,null,35.799999999999997,-3.1189999999999993,38.601677836094538,0.99535294347004244,0.89826156976420057,{"$f":"POWER(10,K25)","v":7.9115498661167969},79.115498661167976]
["Sample 5",1,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0,{"$f":"AVERAGE(M26:M27)","v":0},{"$f":"N26*5","v":0}]
["Sample 5",2,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0]
["Sample 6",1,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0,{"$f":"AVERAGE(M28:M29)","v":0},{"$f":"N28*5","v":0}]
["Sample 6",2,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0]
["Sample 7",1,10,null,null,null,35.022796999999997,-3.1189999999999993,38.601677836094538,0.99535294347004244,1.1474449618770572,{"$f":"POWER(10,K30)","v":14.042517115357439},140.42517115357438,{"$f":"AVERAGE(M30:M31)","v":116.07842529368199},{"$f":"N30*5","v":580.39212646840997}]
["Sample 7",2,10,null,null,null,35.599580000000003,-3.1189999999999993,38.601677836094538,0.99535294347004244,0.96251934469206013,{"$f":"POWER(10,K31)","v":9.1731679433789584},91.73167943378958]
["H2O",1,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0,{"$f":"AVERAGE(M32:M33)","v":0},{"$f":"N32*5","v":0}]
["H2O",2,10,null,null,null,"Undetermined",-3.1189999999999993,38.601677836094538,0.99535294347004244,0,0,0]
[]
[{"$nulls":11},"*One well containes 10ul of templates"]
[{"$nulls":11},"**One elution contains 100ul of templates (extracted from 200ul seminal plasma)"]
[{"$nulls":11},"***Final concentration is calculated per ml seminal plasma"]