Mouse/Rat Growth Hormone ELISA

Contents:

Mel is our resident growth hormone expert

Assay Procedure

Allow all specimens to reach room temperature (~25oC) and mix thoroughly

  • Mark the microtitration strips to be used
  • Prepare the Mouse/Rat GH antibody-conjugate solution by diluting the Mouse/Rat GH Antibody-Enzyme conjugate concentrate in the Assay Buffer
    • This should be done 10-15minutes before use in the assay
    • Mouse/Rat GH antibody-enzyme conjugate concentrate should be diluted at a ration of 1 part into 50 parts of the Assay Buffer
    • If an entire plate is to be used, pipet exactly 240uL of the Mouse/Rat GH Antibody-Enzyme conjugate concentrate and bring up to 12mL with Assay buffer
  • Pipet 50L of the Standards, Controls and unknowns into appropriate wells
  • Add 100uL of the Mouse/Rat GH Antibody-Enzyme conjugate solution into each well
  • Incubate the wells on an orbital microplate shaker at 500-700rpm for 90 minutes at room temperature (~25oC)
  • Aspirate and wash each well 5 times with the wash solution and blot dry by invrting plate on absorbent material
  • Add 100mL of the TMB Chromagen Solution to each well + blank
  • Incubate the wells at 500-700rpm for 15-20minutes at room temperature (~25oC). Avoid exposure to direct sunlight (wrap in foil)
    • Visually monitor the colour change to optimise the incubation time as time will vary
  • Add 100uL of the stopping solution to each well + blank
  • Read the absorbance of the solution in the wells within 30 minutes using a microplate reader set to 450nm
  • Program the zero standard as a “blank”
  • Set the instrument (if possible) to dual wavelength measurement at 450nm with the background wavelength correction set at 600 or 620nm

 

Tests Per Plate

96 Well Plate

6 controls + 1 blank

89 test wells

Therefore approx 44 tests per plate