**[[Preparation before doing procedure for ELISA]]** 
   
=====Procedure for TSH examination=====
  -Dispense 200uL samples in wells of dummy plate according to worklist using 200 uL pipette.
  -Take out the microtiter plate. Don't touch the lower surface of the plate.
  -Dispense 50 uL sample from the dummy plate into the microtiter plate using multichannel pipette.Note the time of starting dispensing of sample. 
  -Take enzyme conjugate in trough.
  -Dispense 50 uL enzyme conjugate in microtiter plate using multichannel pipette.Note time after last column is dispensed with conjugate.
  -Mix the plate in incubator- shaker for 10 sec.
  -Incubate the plate at 24'C in the incubator for 2 hrs from midpoint of start time of sample dispensing and end of conjugate dispensing..
  -After 2 hours , take the plate from the incubator. And wash the plate in the [[ELISA washer]].
  -After washing, Pat dry the plate on tissue paper.
  -Take TMB(Tetramethyl benzedene) substrate in the trough.
  -Dispense 100 uL TMB substrate in the microtiter plates with the multichannel pipette.
  -Mix in incubator cum shaker for 5 sec.
  -Incubate at 24'C in the incubator for 20 minutes.
  -Take stop solution in the trough.
  -Dispense 150 uL stop solution in the microtiter plate.
  -Mix in incubator cum shaker for 5 sec.
  -Incubate for 5 minutes in the incubator at 24'C.
  -Take the plate from incubator and put it in the ELISA reader.
  -Connect ELISA reader with the computer and set Filter i.e at 450 nm . then read plate for 3 times.
  -Take average of the 3 OD . and sort the data and calculate results from the graph derived from Standard.
  -Whenever OD of any sample is more than OD of lowest value of calibrator,result will calculated as "#NA",So release it as "<0.2" microU/ml.
  -Whenever OD of any sample is more than OD of highest value of calibrator,Dilute the sample for 1:10 times in next batch and release the report by multiplying result with 10.
  -If the OD of diluted sample is more than OD of highest value of calibrator,release result as ">value of highest calibrator" e.g.If highest calibrator value is 9.5 microU/ml,then release the report as ">95"microU/ml.  
  -After complition of use of elisa washer,give prime wash with deionised water.



**General procedure for [[ELISA READER]]**

**[[Note on using electronic pipette for ELISA]]**