Lab #12 Report

pdf

School

University Of Arizona *

*We aren’t endorsed by this school

Course

447

Subject

Mechanical Engineering

Date

Jan 9, 2024

Type

pdf

Pages

Uploaded by ChancellorRockJaguar38

McKenna Nichols BME 447 Loeffler 06 December 2023 Chapter 13 Lab Report: Immunosensors Results Task 1: Insulin ELISA Kit No test results for this task. Task 2: Insulin Elisa Kit with Smartphone Camera Insulin Concentration Red Intensity Green Intensity Blue Intensity Absorbance 0 165.758 162.835 147.914 0 7.5 165.566 160.011 91.748 0.2074 15 180.160 175.216 103.526 0.1549 30 177.253 173.063 104.134 0.1524 60 173.544 168.654 100.132 0.1694 90 179.501 173.583 82.886 0.2515 135 180.012 172.044 70.379 0.3226

Task 3: Pregnancy Test (LFA) hCG Concentration Red Intensity Absorbance [Red] Green Intensity Absorbance [Green] Blue Intensity Absorbance [Blue] 0 201 0 197 0 179 0 0.25 155.299 0.1120 124.119 0.2006 123.086 0.1626 0.5 157.956 0.1047 97 0.3077 110 0.2115

Discussion In the first task of this lab, we began by filling 8 wells with 100 HRP. Then to each µ𝐿 well we added 25 of one standard or control. After this, the wells were left to incubate for 60 µ𝐿 minutes at 37 ℃ . We then removed the liquids from the wells and wash each well 3 times with 250 of the wash buffer. We then added 100 of substrate to each well and let the solution µ𝐿 µ𝐿 incubate for 15 minutes in the dark. After incubation, we place 50 of stop solution into each µ𝐿 well. In the second task of this lab, we used our smartphone to take images of each well. Using ImageJ, we obtained the intensity values for all three channels, blue, green, and red. From here we calculated the absorbance values using the blue intensities. We then graphed the blue absorbance versus the insulin concentration. This graph can be seen in the results sections of this report. The red and green intensities showed to stay pretty constant as the insulin concentration changed whereas the blue color intensities decreased as the insulin concentration increased. With the blue intensity significantly lower than the red and green intensities, it is perceived as yellow. This is why we must evaluate the blue intensities to quantify the yellowness of each microplate well. In the third task of this lab, we used three pregnancy test strips and placed them into three solutions with varying hCG concentrations for 3 seconds. After allowing the strips to dry for 5 minutes, we took pictures of each strip. We then used ImageJ to obtain the intensity values for all three channels (red, blue, and green). We then calculated the absorbance for each channel. We then graphed the absorbance versus the concentration of hCG. This graph can be seen in the results section of this lab. Although pregnancy tests are typically analyzed in a positive versus negative manner, they can actually be quantified based on the darkness of the lines. We saw that

Your preview ends here

Eager to read complete document? Join bartleby learn and gain access to the full version

Access to all documents
Unlimited textbook solutions
24/7 expert homework help

as the hCG concentration increased, the absorbance of the color intensity also increased in all three color channels. The red increased at the lowest rate, followed by the blue intensity, and the green increased at the fastest rate. Review Questions 13.3 Why do you have to construct a standard curve each time you make measurements? A standard curve should be constructed each time a measurement is made because it is used to accurately determine the concentration of your sample from the signal generated by an assay. Since the signal is never 100% proportional to the sample, the standard curve can be utilized to correct any small errors so it is clear what concentrations correspond to what values. 13.4 If the smartphone camera processes each pixel at 36-bit (12-bit each for three basic colors), can the slope of standard curve with a cell phone camera be increased (i.e., becomes more sensitive)? Repeat this question if the cell phone camera processes each pixel at 24-bit grayscale. The slope of the standard curve can be increased if the camera processes at 36-bits. This is because it would allow the color code to increase from 0 to 256 ( ) to 0 to 4095 ( 2 8 = 256 ). If the camera processes at 24-bit grayscale, the sensitivity would be increased but 2 12 = 4096 the camera could no longer distinguish between red, green, and blue. Instead it would read everything on the gray scale. 13.5 Compare the standard curves with red, green, and blue pixel intensities. Which color shows the best results, and why? (Hint: use the absorption spectrum of gold nanoparticles—Fig. 13.9).

Comparing the three standard curves shows that the curve for the green pixel intensities has the largest slope and the curve for the red pixel intensities has the smallest slope. When compared to figure 13.9, green also has the largest peak which is an indication of a larger absorbance value. The next largest absorbance is the blue intensity. Pregnancy tests contain anti-hCG- gold nanoparticles which bind to hCG. AuNPs absorb green light which makes red the most noticeable color. This means that red shows the best because it is absorbed less and reflected the most.