Run An ELISA With Controls
Use molecular-test evidence to run an elisa with controls accurately.
- Control logic: Molecular results need positive and negative controls.
- Signal interpretation: Bands, colors, curves, and E-values must be compared to a rule.
Prerequisites are inferred: pending teacher review.
Re-learn the skill with worked practice and clear examples.
Make an ELISA call by comparing the patient's absorbance to the negative and positive controls.
| Well | What is in it | Color reading |
|---|---|---|
| Blank | buffer only | clear (0.0) |
| Negative control | no antibody | clear (0.1) |
| Positive control | known antibody | strong yellow (1.8) |
| Patient sample | patient serum | faint yellow (0.4) |
Use the plate readings. The blank is 0.0, negative control 0.1, positive control 1.8, and the patient sample is 0.4. How should the patient result be read?
Approved| Well | What is in it | Color reading |
|---|---|---|
| Blank | buffer only | clear (0.0) |
| Negative control | no antibody | clear (0.1) |
| Positive control | known antibody | strong yellow (1.8) |
| Patient sample | patient serum | faint yellow (0.4) |
- A.Borderline / weakly positive -- close to the negative control, so retest before calling it positive
- B.Strongly positive, the same as the positive control
- C.Exactly zero, like the blank
- D.Invalid, because a positive control should be clear
Show the worked solution ▾
Answer: A. Borderline / weakly positive -- close to the negative control, so retest before calling it positive
- Step 1: Compare to the controls: Patient 0.4 sits just above the 0.1 negative and far below the 1.8 positive.
- Step 2: Judge the distance: Being much closer to the negative than the positive makes it borderline, not a clear positive.
Why it's right: At 0.4 the patient is near the negative (0.1) and far from the positive (1.8), so it is borderline and should be retested.
- B: 0.4 is far below the 1.8 positive, so it is not strongly positive.
- C: 0.4 is above the blank's 0.0, so it is not zero.
- D: A positive control is supposed to be strong-colored, so 1.8 is correct, not invalid.
Aligned to Biotechnology Research and Experiments · reading level ~grade 9
Use the standard table. The standards read 0.0, 0.5, 1.0, and 1.5 for 0, 10, 20, and 30 units. The patient sample reads 0.5. Using the standards, about how many units does the patient have?
Reviewed| Standard | Absorbance |
|---|---|
| 0 units | 0.0 |
| 10 units | 0.5 |
| 20 units | 1.0 |
| 30 units | 1.5 |
| Patient sample | 0.5 |
- A.About 10 units
- B.About 20 units
- C.About 30 units
- D.0 units
Show the worked solution ▾
Answer: A. About 10 units
- Step 1: Match the reading to a standard: An absorbance of 0.5 lines up with the 10-unit standard, which also reads 0.5.
- Step 2: Read across to the amount: So the patient's 0.5 reading corresponds to about 10 units.
Why it's right: The 10-unit standard reads 0.5, and the patient also reads 0.5, so the patient has about 10 units.
- B: 20 units reads 1.0 on the standards, not 0.5.
- C: 30 units reads 1.5 on the standards, not 0.5.
- D: 0 units reads 0.0; the patient reads 0.5, so it is not zero.
Aligned to Biotechnology Research and Experiments · reading level ~grade 9
- In Unit 1.1 ELISA Lab & Controls, this skill turns class evidence into a result another person can check.
Fill these in as you work through the lesson.
- ELISA (a color-based test for an antibody or antigen):
- Blank (buffer-only well that sets the zero reading):
- Absorbance (the number that goes up as color gets stronger):
- Standard curve (known amounts plotted against their readings):
In an ELISA, a strong color (high absorbance) like the control means the target is present; a clear well like the control means it is absent.
- Which well should read close to zero in a clean run?
- Which well shows what a strong positive looks like?
- If a patient well reads 0.4 between a 0.1 negative and a 1.8 positive, is the target clearly present, absent, or borderline?
Use the plate readings. The blank reads 0.0, negative 0.1, positive 1.8, and the patient 0.4. Compare the patient to the controls and decide whether the result is clearly positive, clearly negative, or borderline, and say why.