Pregnancy Test and ELISA Test Flashcards
Checks your pee or blood for a hormone called human chorionic gonadotropin (hCG)
Pregnancy Test
The body makes this hormone after a fertilized egg attaches to the wall of the uterus
Human Chorionic Gonadotropin (hCG)
The increase of this hormone usually happens about 6 days after fertilization. It rises quickly, doubling every 2 to 3 days
Human Chorionic Gonadotropin
Types of Pregnancy Test
Blood Tests and Urine Tests
You can take these at home or in a doctor’s office. Along with being private and convenient, this type of test is quick and easy to use. It is also very accurate if the directions are followed properly.
Urine Tests/Home Pregnancy Tests
Ways to Perform Urine Tests/Home Pregnancy Tests
- Hold the test stick in your urine stream
- Collect pee in a cup and dip the test stick into it
- Collect pee in a cup and use a dropper to put it into another container
- Wait for a few minutes before seeing the results
- After taking the test, confirm the results with a doctor who can perform more sensitive tests
This type of pregnancy test is performed at the doctor’s office. It can detect pregnancy earlier than a home pregnancy test, about 6 to 8 days after ovulation.
Takes longer to get the results than with a home pregnancy test
Blood Tests
Two Types of Blood Pregnancy Tests
Qualitative hCG Test and Quantitative hCG Test
Simply checks for hCG- It gives a “yes” or “no” answer to the question, “Are you
pregnant?“
• Doctors often order these tests to confirm pregnancy as early as 10 days after conception.
• Some can detect hCG much earlier.
Qualitative hCG Test
measures the exact amount of hCG in your blood.
• It can find even very low levels of hCG.
• These tests may help track problems during pregnancy.
• Your doctor may use them along with other tests to rule out an ectopic pregnancy, when the fertilized
egg implants outside your uterus, or after a miscarriage, when hCG levels fall quickly.
Quantitative hCG Test (beta hCG)
A quantitative immunological procedure in which Ag-Ab reaction is monitored by enzyme measurements. It is highly sensitive
Enzyme-linked Immunosorbent Assay (ELISA)
Samples routinely used in ELISAs
Serum, plasma, cell culture supernates, cell lysates, saliva, tissue lysates, and urine
An ELISA test may be used to diagnose:
HIV, Lyme Disease, Pernicious Anemia, Rotavirus, Syphilis, and others
First to label antibodies with a fluorescent dye, and use it to identify antigens in tissue sections. This method is known today as immunofluorescence.
Albert H. Coons and his colleagues (1941)
Described radioimmunoassay
Rosalyn Sussman and Soloman Berson (1960)
Independently invented the ELISA test
Eva Engvall and Peter Perlman (1971)
A method in which a conjugated substrate competes with a protein of interest. Developed and used to detect hCG
Competitive ELISA Method (1976)
A method in which the detection antibody is coated onto the plate surface before the protein of interest is added; is developed and tested on several substrates for proof-of-concept
Sandwich ELISA Method (1977)
A method in which a secondary antibody is added for detection purposes; developed and used to detect human serum albumin
Indirect ELISA (1978)
What screening test was first used for testing HIV? Give the exact date as well
ELISA Test - March 2, 1985
Discuss the principle behind ELISA
- Uses an enzyme to detect the binding of Ag-Ab
- The enzyme converts a colorless substrate to colored product
- That indicates the presence of Ag
- It can be used to detect the presence of both Ag-Ab
The Four Main Types of ELISAs
- Direct
- Indirect
- Sandwich
- Competitive
When to use Direct ELISA?
Assessing antibody affinity and specificity.
Investigating blocking/inhibitory interactions.
Advantage of Direct ELISA
It is a fast and simple protocol
Disadvantages of Direct ELISA
- Less specific since you are only using 1 antibody.
* Potential for high background if all proteins from a sample are immobilized in well.
When to use Indirect ELISA
Measuring endogenous antibodies
Advantages of Indirect ELISA
- Amplification using a secondary antibody
- Cost Effective
- Different visualization markers can be used with the same primary antibody
Disadvantages of Indirect ELISA
- Potential for cross reactivity caused by secondary antibody
- An extra incubation step is required in the procedure.
When to use Sandwich ELISA
Determining analyte concentration in a biological sample.
When to use Sandwich ELISA
Determining analyte concentration in a biological sample.
When to use Sandwich ELISA
Determining analyte concentration in a biological sample.
Advantages of Sandwich ELISA
- Highest specificity and sensitivity
- Compatible with complex sample matrices
- Flexibility and sensitivity, since both direct and indirect detection methods can be used.
Disadvantages of Sandwich ELISA
- Longer protocol
* Challenging to develop
When to use Competitive ELISA
Determining concentrations of a small molecules and hormones.
Advantages of Competitive ELISA
- Ability to quantitate small molecules
- High specificity, since two antibodies are used.
- High sensitivity, since both direct and indirect detection methods can be used.
- Suitable for complex samples, since the antigen does not require purification prior to measurement.
Disadvantages of Competitive ELISA
- Less specific since you are only using 1 antibody
* Requires a conjugated antigen
The first 4 steps in ELISA
Step 1
Capture antibody binds to ELISA plate wells
Step 2:
Add sample to well antigen within the sample binds to the capture antibody.
Step 3:
Wash microplate Unbound material is washed away, leaving only the antigen of interest
Step 4:
Add detection antibody Enzyme conjugated detection antibody binds to a second site on the antigen of interest
The second 4 steps in ELISA
Step 5:
Wash microplate Unbound antibodies are washed away, leaving only those specific for the target of interest
Step 6:
Add substrate Substrate is converted by the enzyme on the detection antibody, producing a color change
Step 7
Read plate The microplate reader detects the colored reaction product and outputs optical density (OD) values
Step 8:
Calculate results The amount of antigen in each sample is calculated and analyzed
Determines the presence of antigen and/or antibody; Positive or Negative Result
Qualitative Test
Determines the quantity of antibody; No. of positive and negative result
Quantitative Test
• It is a good techniques and being popular today because of its simple and not
involve radiation .
• It is easy than other test .
• It is most useful than other test because of their sensitivity.
• It is not harmful.
ELISA
