Science topic
Parkinson's Disease - Science topic
A topic to discuss Parkinson's disease research: Basic science, clinical research, treatment.
Questions related to Parkinson's Disease
The PD patients I have known never smiled and showed little emotion on the face. Hence I came up with this hypothesis. If the answer is positive, it would be of clinical value.
I am working of LUHMES cells lines. I differentiate them for live cell calcium imaging. On the day of imaging, I stained the with fluo4 in differentiating medium (pH=7.4) and incubate the cells at 37C.
I am acquiring images at 20x, time interval of 1s and sometimes 2s, exposure time (100-150ms) for 30 minutes.
After 5 minutes of acquisition, I stimulate them with 2uM of ATP.
When I analyse the images, I dont observe spikes, in the cells and all the cells behave differently. In most of the cells I dont observe oscillations but increasing concentration of calcium. I also try to do imaging in saline buffer and dpbs but apparently cells did not like it and were stressed even before starting imaging.
Good morning, working on patient serums, I noticed that in western blotting tests the IGs give false positives and unspecific signals. Ask if anyone can give me a methodology to remove these immunoglobulins, considering I have few serum available.
Thank you.
Hello, could anyone help me with the protocol of deparaffinization, rehydration, and antigen retrieval for human brain tissue?
Hi everyone,
I have been having some issues with SHSY5Y cells that I never had in the past. I used to culture them in DMEM:F12 with 10% FBS and 1%Antibiotic-Antimycotic and they grew just fine. Back in October, when we received a new batch of medium, I started seeing increased cell death after 1-2 passages: specifically, I would see debris like structures and the cells literally peeling off from the bottom of the flask. I initially attributed this to the quality of the medium and ruled out incubator, FBS etc issues. I thawed a new vial of cells and I had the same issue. Then I was told to try OptiMEM and with a new vial from ATCC I noticed that they grew beautifully and reached confluence within 3 days. After the 3rd passage, I started seeing the same type of pattern that I saw before--cells fragmenting, a lot of debris and cells peeling off and floating in suspension 1-2 days after plating. I am desperate as my lab has wasted many cell stocks and money on trying to figure out what this is. My next step is to test for mycoplasma but I wasn't sure if this is the case considering that we literally just bought a new stock from ATCC and it grew fine until the third passage.The medium doesn't yellow out and it is not yeast contamination. What can be the source of this type of behavior?Any input is greatly appreciated! :)
We are going to overexpress neurotrophic factors in the striatum and substantia nigra region of mice. We are interested to have overexpression in both neurons and glia. The viruses are intended to be administered sterotactically.
On the RG, I came across two very rarely talked but very important topics.
Abstract TP165: Associations Between Parkinson's Disease and Stroke by @Benjamin Kummer et al 2017 and Risk of stroke in patients with idiopathic Parkinson's Disease by @Claudia Becker et al 2009.
There is not much research available on these hidden topics...
Let me open the stage to unfold scientific research ideas around the world.
Topic of discussion - Does stroke or PD, have a higher chance of causing another disease!! Pharmacological models are available for their research.
For reference -
The importance of disease classification studies for the development of diagnosis & prognosis systems is crucial. Here the main problem is the lack of available medical datasets to use.
For EMG diagnosis, many studies used EMGLab datasets, but the website is NOT available currently.
And I can't find any open-access EMG diagnosis dataset. Parkinson's, SMA, ALS, or other. There are only hand gesture detection datasets present.
Other options are
1- Using an EMG simulation program like EMG-GAN (I don't know how efficient and representative it is)
2- Long-term clinical study with a budget
3- Somehow finding a -retrospective- signal record from a hospital (I don't know how to reach them)
Additionally, I want to know the usability of this short-term prototyping approach :
Simulation of signals by mimicking the patient movements, captured by sEMG device purchased. Can this, highly questionable and insufficient method, be used for the first analysis?
I want to hear your suggestions, thoughts, and shares about the topic :)
Parkinson Disease (PD) is a degenerative disease that affects motor function and sequential.
At which level of H&Y stages will this improve?
Short term or long term effect?
Any evidence to help patient and client?
I have UNM dataset for Parkinson's disease which is publicly available in .mat format. I'm trying to work on it using MNE python and for that I need to get the spatial distribution of the electrodes to generate MNE object.
so far after using scipi.io to read the data I've got following format for each electrode
['FC5']
[]
[[-69.332]]
[[0.40823333]]
[[28.76282344]]
[[76.24736451]]
[[24.16690699]]
[[69.332]]
[[16.518]]
[[85]]
[[6]]
[]
about last two entries 85 is same in all 63 electrodes and 6 here is electrode index but what all these other numbers supposed to mean? where are the coordinates? Can someone explain?
I am working on a project that requires the measurement of neuromelanin purity, and I have found papers that say that they have measured the purity, but they do not say how ( ).
Neuromelanin does not have a commercially available 100% standard, so I cannot do typical comparisons. Does anyone know where to begin or how to do the purity measurements?
Any help would be appreciated.
Thank you!
Dear community,
For my master's project, I want to design an experiment that will asses the efficiency of an antioxidant to stop/restore mitochondrial oxidative stress/redox balance in a PD mouse model, in both early and later stages of the disease. Redox balance will be assessed by using a roGFP lifetime measurements in vivo.
I was thinking to use MPTP model, due to it's effect on the mitochondria, simplicity and practicality. However I heard that this model can be difficult and dangerous to work with and might kill the mice faster than the experiment will end.
There is 6-ohda model, but I was reticent to use it because it has to be injected into the brain directly (which I believe might interfere with my fluorescence measurements) and the mechanisms behind it are not fully understood.
I would highly appreciate if anyone could share their experiences about working with these(or maybe other) models and give me an appreciation of their suitability in regard to my research question.
Thank you!
Hi everyone,
I am currently trying to screen the behavioral effects of the 6-OHDA unilateral injection in the striatum of Wistar male rats to confirm a Parkinson's animal model by apomorphine-induced turning behavior test (or rotameter test).
Although the dose of apomorphine is prepared according to the articles and is injected intraperitoneally to the rats, but instead of turning against the injection site of the neurotoxin, the rats become extremely relaxed and even fall asleep, and it is not possible for me to check the behavior.
I have request from dear students, professors and scientists that help me to find out what the main problem is and share the tips of rotameter test with me.
Please feel free to have contact with me.
Thank you for reading and helping!
Does anyone know what is the first paper published about physical therapy for Parkinson's disease in the world? If so, can share the reference (or the pdf file)?
I appreciate if you could help me!
Hello everyone,
I am using retinoic acid to differentiate SH-SY5Y cells into neuronal cells to generate a Parkinson's disease model. I will perform MTT assay before using a neurotoxin of interest to generate a PD model again for another set of experiments but I am not sure if I have to differentiate the cells again because I have seen that the same cell line is used undifferentiated as well, for example in Alzheimer's disease research.
Is it a "must" for in vitro PD model?
Thank you
Dear Researchers,
What is the best way to deal with imbalanced Dataset when the Imbalanced ratio , IR is more than 4.5 (majority 2000, minority 400) ? If I want to use deep learning methods or transfer learning for classification, what is the best way to augment the data. In my problem Healthy control dataset is the minority.
Hello everyone! I am curious as to whether we can induce mitophagy in HEK293T (or HEK293, which should be similar?) cells, without Parkin transfection?
I have read some papers in the mitophagy field and got an impression that many of them use HeLa cells, which don't express Parkin endogenously, as the model cell line to study mitophagy. For mitophagy induction, they needed to transfect Parkin into the cells and exert chemical treatments afterward. And I was confused why they use HeLa cells to begin with.
If anyone has had success in studying Parkin/Pink1 mitophagy in HEK293T cells, could I get some advice as to what chemicals you use and the duration of the treatment to induce mitophagy? Thanks very much!
I start learning about synchronization between different states and criticality in Parkinson's disease. Would it be possible for someone to recommend a relevant and practical review paper that examines the new challenges arising in this field?
I am working on EEG self-detection of Parkinson's disease and I would need to apply my methods on data from people with Parkinson's disease subjected to ERPs or ssEVPs with a control group of healthy people subjected to the same stimulation under the same conditions.
I have been searching for a year without being able to find a database. So I am looking for a direct link to a laboratory or researcher who can provide me with this data.
Thank you.
I'm studying with a-syn preformed fibril for obtaining Parkinson's Disease model in rodents. We are trying to probe-sonicate the fibril at room temperature using different settings to obtain a fibril fragment within 40-50 nm. So, we tried to adapt your protocol (Generation of Alpha-synuclein Preformed Fibrils from Monomers and Use In Vivo) for our sonication trials; however, we couldn't reach the desired smaller diameter.
Details of our trials:
Trial 1: We used 20-25% amplitude (power of our sonicator 70W and diameter of probe is 3 mm), total 30 sec (1 sec on, 1 sec off) for 5ug/ul PFF solution, but according to the zetasizer result, we obtained fibril fragments of 505 nm average size. In the second step, we extended the time, making it 2 minute. But again, the fibril fragments were average 930 nm. Finally, we diluted the solution to 2 ug/ul from 5 ug/ul, we tried %20-25 amplitude (power of our sonicator 70W), total 60 sec (1 sec on, 1 sec off). Zetasizer results were 470 nm average size.
We thought our sonicator is weaker than in the protocol, so we tried to use more powerful a sonicator than previous at trial 2.
Trial 2: We used %30 amplitude (power of our sonicator 500W and diameter of probe is 2 mm), total 60 sec (1 sec on, 1 sec off) in 5 ug/ul and 2 ug/ul PFF solution. Zetasizer results were 549 nm average size. After then, we continued to sonicate for 2 more minutes (the total 3 minute – 1 sec on 1 sec off). Measurement results were 918 nm as average.
If possible, could you provide us with any suggestions to sonicate the fibrils?
Thanks for your helps.
8 µg (in 1.6 µL) of 6-OHDA was introduced into the right striatum of mice according to the coordinates A = 0.5, L = 2.0, and V = -3.5 relative to bregma. The needle was withdrawn 5 min after the injection. Seven days after operation, 0.5 mg / kg apomorphine was used to induce rotation, Unfortunately, only a few mice were able to induce seven rotations per minute. Can someone please tell me what went wrong? How can I improve the surgical or experimental protocol?
We have published today a paper at Springer Nature journal of Scientific Reports entitled "Optimal time lags from causal prediction model help stratify and forecast nervous system pathology" by Theodoros Bermperidis and co-authors from my lab at Rutgers, colleagues at Stevens Institute of Technology and Columbia University. The paper describes how to standardize signals from wearables and make causal predictions forecasting disorders of genetic origins and unknown etiology. These included Fragile X and SHANK3 deletion syndromes, both linked to Autism, and Parkinson's disease and Vestibular Dizziness Handicap. I wonder if we could use gait more often in the clinical settings now that we have off-the-shelf wearables at our disposal. Research shows that it would improve classification and help stratify heterogeneous disorders like Autism and Parkinson's disease. Read more here https://www.nature.com/articles/s41598-021-00156-2
I see that many papers include analyzing calcium in their methods when the differentiation of iPSCs to neurons is taking place. Why is this necessary?
Conversely, rotenone, which is an insecticide, has.
Adverse health effects from Rotenone have been well documented in the time since the NOSB reviewed botanicals in 1994. But still Rotenone is continue to be used for organic production. Although no longer offered for sale in the USA, rotenone is still in use as a pesticide on organic farms. Among the compelling reasons to prohibit rotenone, the most heavy is it's linked to eye problems and Parkinson's disease. Astornishelly enough is to note that Rotenone's synergist, piperonyl butoxide, is prohibited from use in organic agriculture.
How to make a Parkinson's disease (PD) model of Zebrafish ?
Is there any detail protocol ?
I am trying to figure out how to differentiate between the pars compacta and pars reticulata. In healthy patients you can easily see the difference between the both where pars compacta has a lot of dopaminergic cells and pars reticulata doesn't. However when I look at the SN of Parkinson's patients, the clear/obvious/visible border isn't there anymore. The cells in the Substantia Nigra pars compacta (especially the ventral group, so closest to the pars reticulata) are degenerated. Thus, the clear border is gone. I am not sure where the pars compacta stops now and where the pars reticulata starts.
Are there more hallmarks to define the border between the pars compacta and reticulata?
Is the motor complications in Parkinson's disease are due to early use of L-Dopa or due to progression of the disease
Hello! I'm a Physical Therapy student and currently working on research about stabilizing spoons and their effects on people with PD and ET. I was hoping that some of you may have known any systematic review or RCT articles that I could use as a reference for my study?
Purely asking out of curiosity - we have another protocol that seems successful, so I'm just curious chemically speaking why this might've happened with our first protocol. We initially used a mobile phase of pH 3.0 (MDTM Type II Mobile Phase), and a 2-channel analytical cell set to -100mV and +250mV, where peaks are quantified from the +250mV electrode chromatograph. Despite being the same concentration, HVA peaks are always CONSIDERABLY shorter than all of the other monoamines and metabolites, and consequently detection cuts off at a concentration about 10-fold greater than the other neurotransmitters.
Given the pKa of homovanillic acid (4.4), compared to NE(9.5), DOPAC(3.6), DA(8.93), 5-HIAA(4.22), and 5-HT(9.31), they should all be fully protonated at a pH of 3.0 when suspended in our mobile phase, so I'm not sure why HVA shows such unique differences from the others? Our recording electrode is set to a positive cell potential, so it should be oxidizing (taking electrons from) the sample - could it be that HVA has less 'available' electrons? If so, why is that?
We have supplies coming to switch to a different mobile phase, install a conditioning cell and tackle this problem, but I'm curious to hear why mechanistically this may be happening? Also, again out of curiosity, does anyone know what about a conditioning cell (installed between the column and analytical cell) makes it capable of increasing sensitivity?
I am doing research for Parkinson's disease gait analysis. Where can I find a database based on the acceleration of gait from Parkinson's disease patients?
Patient with neurological problems have different problems. This will impaired their physical, cognitive and even social interaction and so forth.
So, which approach and why that approach is used?
Look into more discussion and information.
I am currently working with SY5Y cell line and my goal is to differentiate them into dopaminergic-like neurons for investigations related to Parkinson`s disease. As one of the markers of dopaminergic neurons I have considered using D1 receptor, but I haven`t been able to identify a single study that expressed this type of dopamine receptors in SY5Y cell line by chemical treatment (e.g. retionic acid or TPA) Many did prove expression of D2 and D3 receptors. So I am wondering should I use D1 receptor at all as a marker of dopaminergic neurons?
We would like to roughtly assess the potential neuroprotective ability of a certain agent to prevent people from developing Parkinson's disease (PD). PD has an incidence of 1% in the general population over 60 years. We are planning to conduct a retrospective study on a group of people, who have all been exposed to this agent for many years. The potential beneficial effect of the agent would also consider information about time of exposure to the agent and the estimated total received dosage of the agent during the lifespan. In case if the agent has neuroprotective properties, we predict people who were exposed to it for a longer period at higher dosages have lower overall prevalence of PD than the general population of the same age group. Our concern is - what would be a minimum number of participants who are 60 years old or older, to be able to estimate the agent's potential beneficial effect?
If I'm right, dopaminergic neurons in substantia nigra but not in ventral tegmental area are affected by Parkinson's disease. I have stained brain slices with TH to observe changes in dopaminergic neurons in Parkinson's mouse model. In the confocal images obtained after staining, I could not demarcate dopaminergic neurons belonging to the substantia nigra and ventral tegmental area. I'll appreciate it if you could suggest to me the way to find the TH-labeled neurons affected in Parkinson's diseases. I have attached images I obtained and I use ImageJ for TH positive cell counting.
Thank you
Saroj
Environmental toxicants induces PD in animal models, one of which is rotenone. Controversies exist on whether the stereotaxic technique or systemic administration is the best. While some have argued that there is high mortality using the systemic method compared to the stereotaxic surgery method. Which method best replicate the pathological hallmarks of PD?
I'm a retired research chemist/physicist with Parkinson's disease and I've been working on upregulating Nrf2 with sulforaphane to fight oxidative stress.
The results I achieved on my PD and with a group of 8 people with PD have shown that sulforaphane strongly attenuated non-motor symptoms especially fatigue and lack of motivation, suggesting that the first target for Nrf2 seems to be mitochondria. I am not an expert in this subject and I would like to reach out to research groups and Parkinson's disease specialists to discuss how to take this idea further.
You can find the background here :
And a preprint I have written on RG here:
Hi,
I'm working on poison induced Parkinson in rats and I do apply some behavioral tests to check Neuropathic pain.
As a result, I find out that they simply show signs of neuropathic pain ( increased mechanical allodynia meaningfully) exactly on the same side I applied lesion to VTA, Substantia nigra and striatum through stereotaxic surgery.
Knowing that there is decussation (crossings in the form of an X, relate each side of the brain to the opposite side of the body.) As a physiological normal situation.
Does anyone have the same experience?
Or Does anyone can explain this complicated result?
I assure you that I have done surgeries and behavioral tests with certitude.
I am trying to assess spatial memory in MPTP induced Parkinson's disease C57Bl6 mice using a Morris water maze. I wanted to know if there are any specific dimensions to the MWM to be used. Most papers suggest using 120 cm diameter with 40 cm height . Has anyone used a maze smaller in diameter than this?
I have tried administering rotenone intraperitoneally to C57Bl/6 mice in my lab, but there is high mortality rate. Subcutaneous rotenone at a dose of 2.5 mg/kg for 28 days did not show any changes in alpha synuclein levels. I m in need of an acute rotenone model of PD, in which I can check the effect of molecule against alpha synuclein aggregation.
I have some files about PET-DAT in patients, having parkinson disease. And I use spm12 implemented in matlab15b for coregistration and spatial normalization of images and voxel-based comparison. But I do not know how to measure regional uptake values for region of interests, like putamen, caudate.
Can you offer some methods and Suggestions?Thank you very much for your help.
Hello Research Gate community,
we are currently trying to differentiate SH-SY5Y cells into dopaminergic neurons. However, every protocol we tried so far did not work out. We tried to induce directed differentiation using RA for 5 days following supplementation with TPA for 5 days in various concentrations (as recommended in several papers). We tried differentiating using RA for 5 days followed by 5 days of BDNF treatment, which also did not work out. Are there more things we have to keep in mind as for example low glucose media or supplementation with Glutamine? We usually use standard DMEM with FCS and Pen/Strep. We would be very happy for any advice.
Thanks a lot for your suggestions.
Current genomics tools to to discover new or study current biomarkers of neurodegenerative diseases.
Hi Jung-Min Lee, sorry to bother you, but we are developing a project to study Physical Activity in Parkinson Disease patients, but we can not download the specific software to work with the sensors. So, can you ask the person who worked for the Sensewear Armband how is it possible to download the software needed to work with the system (sensors)?
If you intend to put me in contact with that person my email is
Thank you.
Best regards, Filipe Melo.
Patient diagnosed with intractable major depression. Olanzapine treatment caused severe drug-induced Parkinsonian symptoms. Olanzapine discontinued 18 months ago and patient recovered from DIP. What alternative medication would you recommend?
We investigate the mechanisms of Parkinson's disease and do drug design for Parkinson's disease. We use theoretical physical chemistry, biophysics and bioinformatics in our studies. We are in need of experimental collaborators to apply for funding from the Michael J Fox foundationand COST EU projects. Are you interested?
One of the current Models prepared to cure Parkinson's disease is based on the process to derive Brain cells from Skin Cells. Is the same possible?
Does anyone have a tip/hint on how to deliver physical assessment remotely? References are welcome! I am working on a project to deliver physical exercises remotely to older people and people with Parkinson's disease, however the research team would like to assess participants' progress too. #COVIDー19
Sleep problems can be seen in patients with Parkinson's disease. Can it be about serotonin?
In mice, ablation of the raphe and no production serotonin increases wakefulness and impairs the homeostatic response to sleep deprivation.(DOI:https://doi.org/10.1016/j.neuron.2019.05.038)
Even in the absence of depression, the CSF levels of 5-I-HAA of patients with Parkinson’s disease are lower than those of age-matched controls.
( DOI: 10.1176/jnp.2.1.88 )
since apomorphine is not availble, and pramipexole and apomorphine are both dopamine receptor agonists, so I'm wondering if I can use other dopamine receptor agonists to verify the rotation test of 6-OHDA induced PD mouse model.
Why is running in Parkinson's disease both possible and have therapeutic effects in contradistinction to walking?
Hi Everyone. I am trying to check the neuroprotective activity of some compounds in the rotenone model of Parkinson's disease in PC-12, for which I will check the effect of that compound on cell viability first. I want to ask whether it is required to differentiate PC-12 for this purpose, as I tried to differentiate the cells using 30 ng/mL as well as 50 ng/mL NGF. I am able to see the differentiated cells in 3-4 days in 96 well plate, but NGF itself is causing cell death, which could be a variable for the cell viability test.
The association of free radicals in the pathophysiology of chronic diseases like degenerative brain disorders (AD, PD, HD, Stroke) has been evident by a substantial research.
We are seeking to identify the major barriers Parkinson's disease patients face in obtaining care for mental health issues. What are the major gaps in treating mental health disturbances in this population?
Hello,
I'm currently designing a protocol in Parkinson's disease and I was wondering about the optimal timing to test motor symptoms in relation to last dose of dopamine agonist(s).
It is clear to me that standardisation of this aspect is essential, but I could not find specific literature with recommendations in this regard.
Any pointers to papers or opinions on this matter would be immensely appreciated.
Thank you
I want to test the effect of a neuroprotective drug on rotenone rat model of Parkinson's disease but after reading the related articles about my work, I became doubts about select the best dosage of the drug and treatment methods. in some of the articles they using a low dosage of a neuroprotective drug and in some high dosage and their treatment was different in some cases, they use the oral method and some intraperitoneal injection method.
Does anyone have any suggestions to select the best dosage and treatment (o.p or i.p)?
Many thanks
Can histone deacetylase activity influence the progression of Parkinson disease? If so, what are the underlying mechanisms?
Current researches to cure Parkinson's disease deal with the formation of the dopaminergic neurons from the Skin fibroblasts but even after the formation of neurons, is their administration to the Hippocampus region of Brain possible?
If yes, is that economically feasible?
Parkinson's Disease is a seerious issue now days.
Yoga may be a way of manage this problem. What yoga practice should be provided to them?
I have found only a handful of studies in my review of literature. Just wondering if anyone is looking at this also.
According to researches, appendix might has connection with Parkinson's Disease but consequences are conflict. One study says, appendix removal reduces risk of PD but another study says that it increases.
Uncovering a Link Between the Appendix and Parkinson Disease Risk
doi:10.1001/jama.2019.9041
Thank you!
I'm looking for the biochemical explanation behind the creation of genetically modified organisms (GMOs) for use in research of human pathologies such as Parkinson's Disease (PD) or Huntington's Disease (HD). I will really appreciate any answer or recommendation of literature.
I'm interested to continue with brain signal research in case parkinson disease. I'm searching for EEG data for the same or interested to work with the research groups in the same field.
Hi, we're working on a pilot research and our aim is to develope parkinson dis. with both intranuclear and intraperitoneal injection methods.
In i.p. model The problem is rats can not tolerate the protocol recommended in articles [ examined dosages : 2-2.5 mg/kg dissolved in DMSO and diluted in MCT i.p. injection ]. We found Rats dead after 2-3 consecutive days.
What's the solution ?
Need to consider all variability arising from age, gender, post mortem interval etc.
Here is the latest example of our 20 years of cognitive dissonance. That the "heterogeneity of PD" remains "a major problem" but that better techniques (such as "utilization of complex analysis strategies and automated standardized assessment methods") will circumvent it. If even LRRK2-PD, as indirectly suggested by the authors, represents a collection of different diseases, can we rely on the brute force of newer and sophisticated analytic methods to bring us out of the conundrum that our unabated allegiance to PD as a unitary disease construct brings? Our field remains stuck in the quest to validate the century-old clinico-pathologic, convergent model of disease nosology, unlike most other fields of medicine, ahead by three decades of embracing systems biology divergence. No analytic technique will overcome the basic flaw in our ongoing biomarker-discovery efforts: that we, clinicians, get to set the gold standard against which to validate biology. It should be the other way around: biology validating disease subtypes. Unless we start walking the talk of PD as a syndrome of many biologically distinct diseases --we will continue banging our heads against the same wall. It is time to move beyond the search for biomarkers of convergence. Whatever is found to be common to all forms of PD is unlikely to be pathogenic in any.
I wanted to work on developing a deep Learning model to differentiate different classes of disease and controls. But I am not able to find the required database for it. Can someone suggest me where I can find one?
Or can anyone tell me how do I look for database related to particular icd_9 codes?
We already know the application of botilinum toxine drooling in patients with Parkinson's disease. We often recommend that you swallow saliva and chew gum. What are the methods you use in your daily practice?
Parkinson's disease (PD) is a neurodegenerative disorder that affects predominately dopaminergic neurons in a specific area of the brain called substantia nigra. While Parkinson’s itself is not fatal, disease complications can be serious. The Centers for Disease Control and Prevention (CDC) rated complications from PD as the 14th cause of death in the United States.
Currently, all therapies used for PD improve symptoms without slowing or halting the disease progression.
So at present what is the life expectancy of a Parkinson's patient?
This is a very and unique occasion to partecipate at the first convention in which patients, caregivers and Doctors will work toghether in a brain storming....

Hi,
I've been doing several western blot trying to check protein expression, oligomer formation and protein degradation in alpha synuclein in cell lines (SH, PC12 and HEK) after transfection with plasmids (pIres).
I've had a lot of varibiality in my results and I don't know what I'm doing wrong, or why I have such variability in my blots that make them non consistent.
I don't use native gels, but I still see monomers, dimers, trimers and tetramers of alpha synuclein.
I've tested Old and fresh RIPA buffer. I've also tested membrane fixation with 0.4% PFA which improve a bit the bands.
Sometimes I get monomers and dimers for SH, and dimers, trimers, and tetramers for PC12, and sometimes only monomers in SH, and only tetramers for PC12.
I've get other weird results like no tubulin in PC12, or only GFP in the lane where is the C- with pIres emtpy vector only expressing GFP, but not in the rest of lanes with pIres expressing alpha synuclein and GFP (and I confirm that cells were green).
So, I'm desperate and I will appreciate all your help, please!
Attached the procedure and also some images.
Thanks!
The detailed protocol is:
Cells in 24 well plate as C- or transfected with pIres plasmid with alpha synuclein-6xHis tag and GFP
- Remove medium
- Trypsinize and inactivate. Collect in a eppendorf and add 500 ul of PBS to wash them
- Centrifuge 5 min 300 g RT
- Aspirate the SN and freeze the pellet in dry ice. Store the samples at -80ºC
- Lysis buffer: RIPA (50 mM Tris-HCl pH 7,5; 1% TritonX100; 1% NaDeoxycholate; 150 mM NaCl; 1 mM EDTA; 10 mM NaF) + 100x Halt Inhibitor
- Add 100 ul per sample of lysis buffer and vortex until pellet get resuspended. Incubate 30 min on ice and vortex at minute 15.
- Centrifuge 10 min at 4ºC 10.000 rpm
- Transfer the supernatant to a new epp and store the SN and the pellet at -80ºC until use.
- BCA quantification using 10 ul of sample
Western blot:
- 5-10 ug of sample + 4x loading buffer (10% b-mercaptoethanol), 95ºC 10 min + spin
- Load the samples on Precast Gel TGX Biorad (4-20%) Electrophoresis: 100 v 1h 30 min
- Open the cassete in transfer buffer and place the gel over the PVDF membrane (BIORAD already prepare for Tran Blot Turbo System (semy dry system). I usually soak the transfer cuvette, filters and membrane and remove the excess of buffer. Transfer of a mini (2.5A cte up to 25 V 3 min) or a midi or 2 gels (2.5 A cte up to 25, 7 min) gel to PVDF membrane
- 1x TBS wash
- 0,4% PFA fixation 30 min RT
- 2x TBS wash
- Ponceau staining 5 min
- 3x TBS-T wash
- Blocking 5% milk in TBS-T (1-2 h RT)
- Primary antibody rabbit anti alpha synuclein (PA5-17820) 1:1000 in TBS-T ON at 4ºC
- 3x TBS-T wash
- Secondary antibody goat anti rabbit –HRP 1:40.000 1-2 h RT
- 3x TBS-T wash
- Dry excess and put in a dry tray
- Develop with SuperSignal West Pico PLUS Chemilluminiscence (2 ml of mixed buffers and incubate 5 min).
- Remove the excess of substrate and place it between transparent plastic
- Imaging with Amersham 600RGB (1sec, 1 min and incremental ever 1 or 3 or 5 min....)



Why taste sensation is altered by levadopa?
I am started studying on Parkinson's disease, which is neurological disorder...is their any characterization should i follow to study and their prevention.