could you please edit this part of my report? 1. Materials and methods 2.1 Expression of MFE-23 in E.coli. Three kinds of 200 µl bacterial culture with MFE-23 (His and/or Myc tagged) were inocul

fiona

could you please edit this part of my report? 1. Materials and methods 2.1 Expression of MFE-23 in E.coli. Three kinds of 200 µl bacterial culture with MFE-23 (His and/or Myc tagged) were inoculated into corresponding tube respectively and were put into the Orbital Shaker at 37˚C and 225 rpm. The culture with growth media (2TY), ampicilin and 0.5% glucose served as negative control. A spectrophotometer at 600nm was used to measure and record the optical density (OD) at 30 min intervals until OD600≅0.9 and growth curves were established. The addition of 20 µl of 1M isopropyl-β-D-1-thiogalactopyranoside (IPTG, Sigma) to the cultures was used to induce the expression of MFE-23 when OD600≅0.9. After induction, the cultures were placed back to the shaker for incubation overnight at 30˚C and 225 rpm. On the next day, the benchtop centrifuge was set up at 4000 rpm for 20 minutes for the acquirement of bacterial supernatants and the supernatants (sample 1, 2, 3) moved to Falcon tubes as a preperation for ELISA experiment． 1.2 Identification of tagged MFE-23 antibody Half of the 96-well ELISA plates were coated with 100 µl of the purified CEA (1 µg/ml) for each well, while each of the wells was treated with the same amount of phosphate-buffered saline (PBS). The plate with plastic film was incubated for 1h at RT, and subsequently the wells were washed twice with PBS. The plate was blocked through the incubation of 5% solution (Marvel milk/PBS) with 200 µl for each well at 4˚C overnight. After the washing step was repeated, 100 µl/well of the collected supernatants from bacterial cultures (sample 1, 2, 3), and also the culture (2TY) as negative control were added in triplicate wells to the CEA-coated plates. The plates were incubated for 1h at RT before washed with 0.1% PBS/T twice and then with PBS twice. Three kinds of primary antibodies, including the polysera anti-MFE-23, monoclonal anti-His (Sigma) and anti-Myc (Sigma) were used to capture the tagged scFv and all of them were in a 1:1000 dilution in 1% blocking solution. Every kind of antibodies (100 µl/well) was applied to the wells with every sample, control (2TY) and also the PBS-coated wells. After the plate was incubated for 1h at RT, the washing step was repeated.

Dec 2, 2015 12:03 AM

Corrections · 1

could you please edit review this part of my report?

1. Materials and methods

2.1 Expression of MFE-23 in E.coli.

Three kinds of 200 µl bacterial culture with MFE-23 (His and/or Myc tagged) were inoculated into corresponding tube respectively the respective tubes and were put into the Orbital Shaker at 37˚C and 225 rpm.

You cannot write three 'kinds' - the specific contents of the culture is needed. The written English is not wrong but not precise enough for scientific English.

The culture with growth media (2TY), ampicillin and 0.5% glucose served as negative control. A spectrophotometer at 600nm was used to measure and record the optical density (OD) at 30 min intervals until OD600≅0.9 and growth curves were established.

The addition of 20 µl of 1M isopropyl-β-D-1-thiogalactopyranoside (IPTG, Sigma) to the cultures was used to induce the expression of MFE-23 when OD600≅0.9.

Was the thiogalactopyranoside added to all cultures including the negative control?

After induction, the cultures were placed back into the shaker Orbital Shaker for incubation overnight at 30˚C and 225 rpm. On the next day, the benchtop centrifuge was set up at 4000 rpm for 20 minutes for the acquirement of bacterial supernatants and the supernatants (sample 1, 2, 3) moved to Falcon tubes as a preperation for ELISA experiment．

1.2 Identification of tagged MFE-23 antibody

Half of the 96-well ELISA plates were coated with 100 µl of the purified CEA (1 µg/ml) for each well, while each of the wells was treated with the same amount of phosphate-buffered saline (PBS).

Not clear. Half of the wells had PBS or all of the wells had PBS?

The plate with plastic film was incubated for 1h at RT, and subsequently the (all or half?) wells were washed twice with PBS. The plate was blocked through the incubation of 5% solution (Marvel milk/PBS) with 200 µl for each well at 4˚C overnight. After the washing step was repeated, 100 µl/well of the collected supernatants from bacterial cultures (sample 1, 2, 3), and also the culture (2TY) as negative control were added in triplicate wells to the CEA-coated plates. The plates were incubated for 1h at RT before washed with 0.1% PBS/T twice and then with PBS twice.

Three kinds of primary antibodies, including the polysera anti-MFE-23, monoclonal anti-His (Sigma) and anti-Myc (Sigma) were used to capture the tagged scFv and all of them were in a 1:1000 dilution in 1% blocking solution. Every kind of antibodies (100 µl/well) was applied to the wells with every sample, control (2TY) and also the PBS-coated wells.

"Every kind" is not a good phrase to use. Better to state each kind of antibody to be more precise.

After the plate was incubated for 1h at RT, the washing step was repeated.

It's a pretty good piece of writing. You need to be more precise with some areas so that a person in another lab can repeat your experiment exactly step by step without needing further clarification.

December 2, 2015

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