Lec 4: Hormone quantification

Need to know:

  • Will understand the principle of competitive

assays

• Will be able to discuss how ELISA and RIA assays

work to be able to quantify target analytes

  • Gametes produced in fish (Diagram)

  • Standard curves (Pipetting) - ELISA (Flat of curve) (Recovery rate)

  • Understand this concept Competitive assay.

Endocrine system:

  • Hormones

  • Chemical messengers

Gonadatropins -Chemical messengers telling Gonads what to do.

00genesis and hormines: Bitologenesis

-Hormones (Estrogen), are driving this development

  • Packing oocyte

  • Providing nutrients

  • Maturation - Ovulated eggs

  • Ready for fertilisation

Measure hormones:

  • Antibodies specific to hormone or antrogen of interest.

  • Antibodies will bind to those specific hormones

  • Tracer hormones - Pick up these hormones

Buy kits online, to measure hormones, proteins, vitamins as long as antibodies are specific for the target antigen.

Competitive binding:

ELISA

Enzymes use immune structures, that are binding antrigens of interest, the assay measures.

Estrogen ELISA kit - Colour will react when tracer molecule, reacts. Measure intensity in spectrometer.

Kit comes with estrogen molecule attatched with enzyme.

Plasma sample - Unknown conc

Tracer - Known conc

They compete with a limited number of antibodies they combine too

Spectromtrometer - Measure absorbance accurately, converting absorbacne into a conc of steroid.

Standard curve plot:

Most concentrated tracer - More colour

Radio Immuno Assay:

  • Not anchored

  • Radioactive label, no colour reaction. (Competitiion to attatching to antibodies)

ELISA assay - Low toxicity method.