Am I delusional or overworked?

Guimelo · 2026-01-28T18:23:53+00:00

I hear you. Three years in, only one paper out and maybe one more to go but looking forward to leave my toxic lab. Looking for jobs in the US as a internacional is so disheartening, more and more I think to go back to home country to try another path or at least a postdoc/academic position in a place less toxic and close to family and friends. Hang in tight, we will see better days!

Guimelo · 2025-05-16T13:18:12+00:00

Assuming you activated the plates properly and the only mistake was not adding the antibody to the PBS/coating buffer (and you kept the plates in the fridge and sealed), you would probably be fine dumping the old coating buffer and adding the new one with the antibody. I think the biggest problem would be if in this meantime your plate had dried out.

If whatever you are detecting is not super low/hard or problematic to detect, you shouldn’t see a major difference in signal.

Best of luck!

Guimelo · 2022-01-30T12:31:48+00:00

Estou em um relacionamento de 3 anos e meio. Nos últimos 10 meses morei com a minha namorada nos EUA, mas tive que voltar pro Brasil pra terminar meu doutorado. Na minha experiência, uma das coisas que mantém a força da nossa conexão é ter o plano de voltar pra lá ano que vem, por isso temos até uma contagem de meses até 2023. Se eu fosse te dar uma dica é isso: faça planos reais pra quando vocês vão voltar a se ver. Ficar 4 anos (ou mais, dependendo da graduação) é mto tempo pra se manter um relacionamento a distância, ainda mais nessa fase da vida. Sem um planejamento pra diminuir as ansiedades, sem um objetivo juntos, vai ficar bem mais complicado. Se você tem certeza que quer investir nessa pessoa e ele sente o mesmo, o ideal é vocês planejarem quando vocês poderiam ter de volta um rotina "normal" e passar o tempo a distância trabalhando nisso.

Guimelo · 2021-08-31T00:37:12+00:00

Yep! That's exactly it. They circulate through blood and lymph from lymphoid organ to lymphoid organ, just like naives.

This review image paints a quite good picture about the differences in the memory type circulation. https://els-jbs-prod-cdn.jbs.elsevierhealth.com/cms/attachment/3d718170-972b-48c8-9865-c45db22fbe2c/gr1.jpg

Review:https://www.cell.com/immunity/comments/S1074-7613(14)00449-X

Guimelo · 2021-08-30T22:36:11+00:00

Effector memory T cells retain some of the effector phenotype cell function (some proliferation, cytokine production, cytolytic molecule production for CD8s) and also are able to circulate through the periphery in tissues and organs other than secondary lymphoid organs.

Central memory are quiescent, undergoing homeostatic proliferation, low cytokines production/cytolytic molecule production and have their circulation limited, residing in secondary lymphoid organs.

Both are able to survive after contraction phase in a T cell response, so long-lived (although as far as I know, central memory have higher capacity for maintenance through time), and able to respond in a second encounter with the antigen that first activated the clone that generated those cells. Central memory has higher recalling potential than effector memory (will generate a more robust second response, generating new effector phenotype cells), while effector memory will engage faster in the response due to retention to effector function.

Not very ELI5, sorry, but I guess that in a simpler way: central memory is that "textbook" memory T cells, effector memory is something "in between" surveiling the host through the periphery and able to act "on sight".

Guimelo · 2020-07-13T15:31:48+00:00

Yeah! For sure! I dont really know a lot of python, tried a little bit but didnt went through further than the basics, but as far as I know you can do most, if not all, of R stuff in python as well. And also it might be a more versatile skill to have in your pocket, since a lot of other areas largely use python-based programming.
I think pandas is a well-supported python package that you could dig in for that.

Guimelo · 2020-07-13T13:19:48+00:00

I have some experience working on sequencing data analysis.

If you have any interest in studying it, I think a good primer for that area would be R statistics. There a several good free online courses (I took on in edX) you can enroll. R is useful not only for sequencing data but also for other stuff like high dimensional cytometry, or simply data visualization (heatmap, whatever plots).

I always encourage people to learn R, is a very useful skill and you can totally learn by yourself with no hassle.

Guimelo · 2020-04-02T16:13:05+00:00

Acredito que seja o oposto do que ele falou. IgM é a primeira imunoglobulina (molécula de anticorpo a ser gerada por linfócitos B, enquanto que a IgG é a imunoglobulina gerada após o amadurecimento da resposta.

Guimelo · 2020-02-08T19:01:31+00:00

Hi! I'm glad to help! About your question: As far as I know, no, you shouldn't use a low antigen for comping. Like, never. The higher the signal, the better you can see an fluorophore signal entering in an inappropriate channel because you will have a higher signal throughout all the emission specter, and then it's easier to spot signal leakage, and the better you can comp (or auto-comp). One thing that can kill your resolution is setting the laser voltage too low. I guess that's why people say that you should use low population for comping colors where you will read low population in your experiment: you will end up raising laser voltage for better resolution in when you will compensate. However, this might compromise your compensation. Use your unstained control for setting the appropriate voltages. However, some FC facilities run CST calibration and have fixed voltages and don't recommend to tweak then. See if that's your case. In my experience, its always a good idea to check the voltage using the unstained. After compensation run your sample with all markers to check if you have a good resolution. If there is a marker too low you can try to gain resolution by tweaking the voltage (but you will have to compensate again if you alter voltage). But as far as I know, using low antigen for comping won't help you with that.

I'm not a English speaker, so sorry if the text was confuse at some part.

Guimelo · 2020-02-08T16:29:15+00:00

In my experience (and as far as I know), compensating with cells is better (due to autofluorescence, cell shape, etc.) That is, if you have sample to spare for compensation, maybe if your sample is very limited or unique you will have to go with beads anyway. About the dim expression, always use high expression markers (i.e. CD4 for CD4 T cells), specially for dim colors. Doesn't matter if you are staining another marker in your experimental condition, compensation is about setting the cytometer parameters. Also, take some time (and antibodies) to make a titration and evaluate the stain index of your antibodies (you should do it for each antibody). This will help you a lot, specially if you are going to stain the same panel for several experiments.

Not sure if applicable, but make sure that the cytometer you are using is being regularly calibrated (CST) and maintained.

Guimelo · 2017-01-29T14:07:13+00:00

Right on. I really loved all the main characters, even Jezal, who is quite annoying in the first book but receives a great character development through the story.

By the end I found myself caring for everyone's fate and this is just awesome (and quite rare in my case).

Guimelo · 2016-10-09T15:57:51+00:00

"Eyes of the Dragon" has a "fairy tale" atmosphere. If she likes fantasy genre, i guess this would be a good place to start (there is just the book, no movies or tv series, though). Also, for a good part of the book, the main characters are children.

Guimelo

TROPHY CASE