Difference between revisions of "Part:BBa K2876014:Experience"
| Line 7: | Line 7: | ||
[[File:SCAP2h_protein.png|400px]] | [[File:SCAP2h_protein.png|400px]] | ||
| − | |||
Figure 1: Single Chain Antibody Two Hybrid Protein Detection System | Figure 1: Single Chain Antibody Two Hybrid Protein Detection System | ||
| Line 16: | Line 15: | ||
Figure 2: A graph of the absorbance at 540 nm from an ELISA of DH5-alpha cells transformed to produce IL1B (in orange) compared to the IL1B-Standard (in blue) and the negative control of untransformed DH5-alpha cells (in grey). Protein was extracted using a B-PER protocol. We used the negative control to calculate a null hypothesis for the log of absorbance when there is no protein. H0 Absorbance = Log(0.082) = -1.086. We then did a right-tailed hypothesis test to see if our values of Human Interleukin-1 Beta were statistically significant. The z-score was 21.57; the cutoff for statistical significance was a z-score of 1.895. Therefore, our data was statistically significant and we can accept the alternative hypothesis, Ha: Absorbance =/= -1.806. This leads us to conclude that IL1B protein recognizable by IL1B antibodies is being made. | Figure 2: A graph of the absorbance at 540 nm from an ELISA of DH5-alpha cells transformed to produce IL1B (in orange) compared to the IL1B-Standard (in blue) and the negative control of untransformed DH5-alpha cells (in grey). Protein was extracted using a B-PER protocol. We used the negative control to calculate a null hypothesis for the log of absorbance when there is no protein. H0 Absorbance = Log(0.082) = -1.086. We then did a right-tailed hypothesis test to see if our values of Human Interleukin-1 Beta were statistically significant. The z-score was 21.57; the cutoff for statistical significance was a z-score of 1.895. Therefore, our data was statistically significant and we can accept the alternative hypothesis, Ha: Absorbance =/= -1.806. This leads us to conclude that IL1B protein recognizable by IL1B antibodies is being made. | ||
| + | |||
[[File:IL1B ELISA Pre-stop.jpg|400px]] | [[File:IL1B ELISA Pre-stop.jpg|400px]] | ||
| − | |||
Figure 3: A photo of the IL1B ELISA showing color change as a function of IL1B presence. Columns 1,2, and 6 show the ELISA standards, from least (row A) to most (row H) diluted. Columns 3, 4, and 5 rows A-D are DH5-alpha cells transformed with our IL1B plasmid. Columns 3, 4, and 5 row E are untransformed DH5-alpha cells. | Figure 3: A photo of the IL1B ELISA showing color change as a function of IL1B presence. Columns 1,2, and 6 show the ELISA standards, from least (row A) to most (row H) diluted. Columns 3, 4, and 5 rows A-D are DH5-alpha cells transformed with our IL1B plasmid. Columns 3, 4, and 5 row E are untransformed DH5-alpha cells. | ||
| + | |||
[[File:IL1B ELISApoststop.png|400px]] | [[File:IL1B ELISApoststop.png|400px]] | ||
| − | |||
Figure 4: A photo of the IL1B ELISA showing color change as a function of IL1B presence, after stop-solution added. Columns 1,2, and 6 show the ELISA standards, from least (row A) to most (row H) diluted. Columns 3, 4, and 5 rows A-D are DH5-alpha cells transformed with our IL1B plasmid. Columns 3, 4, and 5 row E are untransformed DH5-alpha cells. | Figure 4: A photo of the IL1B ELISA showing color change as a function of IL1B presence, after stop-solution added. Columns 1,2, and 6 show the ELISA standards, from least (row A) to most (row H) diluted. Columns 3, 4, and 5 rows A-D are DH5-alpha cells transformed with our IL1B plasmid. Columns 3, 4, and 5 row E are untransformed DH5-alpha cells. | ||
Revision as of 01:20, 18 October 2018
This experience page is provided so that any user may enter their experience using this part.
Please enter
how you used this part and how it worked out.
This sequence produces protein interleukin-1B, a human protein produced during fevers. For our project we detected IL1B with two single-chain antibodies fused to transcription initiating proteins (BBa_K2876001, BBa_K2876002). We validated our IL1B with an ELISA (https://www.thermofisher.com/elisa/product/IL-1-beta-Human-ELISA-Kit/BMS224-2) which proved production of IL1B, and that the IL1B produced was folded properly so that antibodies could bind. Validation of antibody binding was essential to ensure that IL1B could function as the target protein in our Single-Chain Antibody Prokaryotic Two Hybrid Detection System (Figure 1).
Figure 1: Single Chain Antibody Two Hybrid Protein Detection System
Figure 2: A graph of the absorbance at 540 nm from an ELISA of DH5-alpha cells transformed to produce IL1B (in orange) compared to the IL1B-Standard (in blue) and the negative control of untransformed DH5-alpha cells (in grey). Protein was extracted using a B-PER protocol. We used the negative control to calculate a null hypothesis for the log of absorbance when there is no protein. H0 Absorbance = Log(0.082) = -1.086. We then did a right-tailed hypothesis test to see if our values of Human Interleukin-1 Beta were statistically significant. The z-score was 21.57; the cutoff for statistical significance was a z-score of 1.895. Therefore, our data was statistically significant and we can accept the alternative hypothesis, Ha: Absorbance =/= -1.806. This leads us to conclude that IL1B protein recognizable by IL1B antibodies is being made.
Figure 3: A photo of the IL1B ELISA showing color change as a function of IL1B presence. Columns 1,2, and 6 show the ELISA standards, from least (row A) to most (row H) diluted. Columns 3, 4, and 5 rows A-D are DH5-alpha cells transformed with our IL1B plasmid. Columns 3, 4, and 5 row E are untransformed DH5-alpha cells.
Figure 4: A photo of the IL1B ELISA showing color change as a function of IL1B presence, after stop-solution added. Columns 1,2, and 6 show the ELISA standards, from least (row A) to most (row H) diluted. Columns 3, 4, and 5 rows A-D are DH5-alpha cells transformed with our IL1B plasmid. Columns 3, 4, and 5 row E are untransformed DH5-alpha cells.
Applications of BBa_K2876014
User Reviews
UNIQ22791bdc74925b5c-partinfo-00000000-QINU UNIQ22791bdc74925b5c-partinfo-00000001-QINU