Agree it's trivially easy to do, but as anyone you does a lot of qPCR knows, they're rife with false positives.

Any environmental contamination will show up as a positive, particularly using the non-quantatative end-point method mentioned above. There's simple controls for that, but still....

> Agree it's trivially easy to do, but as anyone you does a lot of qPCR knows, they're rife with false positives.

Honestly we'd be better off having a widely available test that gives 50% false positives. Worst case, a bunch of people quarantine unnecessarily, but we at least catch all the infected. The alternative is what we have now: widespread community transmission with no end in sight.

With a 50% false positive rate, most of your positives are false positive. You might as well just tell everyone who asks for a test that they're infected.

Having 50% of all tests come back positive is way too high and makes the test basically useless. But having 50% or more of positive tests being false positives is probably the right tradeoff, though you probably want to re-test people who don't show symptoms.

From a public health perspective ... this could be a brilliant solution.

Until they all decide that they’re immune and don’t need to worry about it anymore.

Exactly. I think many people, upon learning they're positive by asymptomatic would (maybe) wait a week to make sure they don't feel ill, and then go out into the world, potentially spreading the virus with abandon because they're among the group that doesn't get ill from it.

You should account for ignorance in public health policy, but you shouldn't rely on it.

Having more false positives than true positives only works if the people taking the test either don't understand statistics or don't know the test accuracy. I for one would probably ignore a positive result if I thought my chances of true positive were 0.001 and my chances of a false positive were 0.5.

That's a great way to lose any remaining confidence the population might have on the establishment. It is also probably unethical so I doubt any doctor would go with it.

It's a perfectly valid concern, but I don't think it needs to turn out the way your fear.

How about we try this: when reporting "facts", how about we try noting that there is uncertainty involved? I know the average Joe and Jane public aren't intellectuals, and it is often how stupid people can be, but on the other hand I think it is also fair to say that we underestimate people.

I think we should at least consider trying this (noting uncertainty in reporting) - it's times like this where we need more ideas and more thinking, not less.

Until Fox News or the governor of West Virginia blows the illusion away.

Huh, I'm lost in this thread.

So, We lie to people and tell em they are infected. And then we are angry that FOX or the governor of WV are going to expose our scam?

What did I miss...

You could tell them the test came back positive. That's not really a lie.

50% is a bit of information. You can repeat the test - even if you consider that some contamination survives repeated tests, if you even get half a bit from subsequent tests you can easily get to eliminate quite a bit of population.

Yea but not nearly enough of the population for this to meaningfully change the response, and now you’ve made the testing results completely meaningless so that we can’t track the disease. Maybe it’s not intuitive to people but we are actually worse off with a test that’s this bad than just the extremely limited testing that’s being done now

I admit it's not intuitive. Tell me more?

I'm assuming there's high false positive rate. If the test has even moderate false negative, then yes it's worse than useless.

High false positive rate (I’m talking like more than 1%) means

(1) you are overwhelmingly unlikely to actually have the disease even if your test comes back positive but you don’t have obvious symptoms

(2) the pool of people testing positive is completely dominated by false positives. The fraction of people testing positive that will actually have the disease would be very small, so you lose the epidemiological benefit of the test (now you can’t actually track the disease well).

So — test is useless for people like you and me who don’t have symptoms or a strong prior probability of having covid BEFORE the test, because even a positive result is overwhelmingly more likely to be wrong, AND now the epidemiologists trying to actually track the disease are instead overwhelmed with false positives so the true signal disappears. It’s bad all around. You could imagine using this as a screening tool, but the false positive rate has to be small enough for that purpose as well (like, less than one percent).

There is a big system-level difference: with 50% false positive, half of the population goes back to living regular lives. False positives stay home, and go to a hospital only if they need a ventialtor (which very few will need since they are not actually sick)

There are 50% false positive, and there are some false negative. And some more not exactly false negative, but simply got sick after the test. And you tell them: we tested everyone, all negatives can just resume normal life. What could go wrong?

If you can't breath you go to the hospital regardless of it being COVID-19.

This is an interesting tradeoff to consider. Under many many circumstances I could see that rate of false positives being awful and intolerable.

In the context of a bona fide pandemic with a massive shortage of desperately needed testing, however, this would be a palatable improvement upon the status quo.

In the case of a generally mild disease, it's much less of a problem. Usually, the problem with false positives is unnessary, risky treatments and overwhelming the medical system. But in this case, you just need to know whether to self quarantine. You only need treatment if symptoms become severe.

Until the global economy unnecessarily collapses.

You also need to consider that when the false positives get over their flu or cold or whatever, they will assume they are immune to COVID.

Really? That test is meaningless. It screws up the numbers, and you might as well do a complete quarantine anyway, since you need to not just quarantine but isolate anyone who’s positive.

Let’s say there’s 100,000 people in the US with covid right now (like 10x the measured number)

FPR =0.5 TPR = 1

Disease rate = 1e5/3.3e8 = 3e-4

P(you have covid| test is positive) = 3e-4/(0.5(1-3e-4) + 3e-4) = 0.06%!!

This is less than useless. Now instead of tracking the disease you are completely overwhelmed with false positives and the result is exactly the same — everyone stay home.

hmm, how many false negatives is it giving?

A 50% false positive rate means that, since most people don't have the virus, about 50% of all the tests will come back positive. You just about might as well flip a coin instead.

This is a classic statistical fallacy. A test that is 50% accurate serves 0 purpose. You might as well consider coin flipping a valid test. If you tested America with this test you’d end up thinking half the country is infected. And within the ones that tested positive only half would actually be and within the negatives half would be and you would have missed them.

A test needs to be materially more accurate than the odds of having the disease to be worth anything.

Classic thought experiment: if 0.0001% of the population has a disease and a test is 99% effective and you test positive, what are the odds you have the disease? Answer: 1%.

Complimentary thought experiment: if an expensive preventive drug was available in limited supply (for 0.1% of the population only) and the earlier you took it the more preventive it was should you give it blindly to as many positives as you can? Probably not because 99% of that would be going to waste. And you would run out of drugs to cover the actual sick. Only 10% of the sick would end up actually getting the drugs.

Isn't the false positive rate the percentage of positives that should have been negative? E.g. if I test 10000 people and 100 of them test positive, but really only 50 of those are sick (and almost none of the other 9900 people are), then I have a false positive rate of 50% and a very useful test. Obviously, this is very different from flipping a coin. And flipping a coin could also have a very high false negative rate.

It's the rate of actual negatives that are perceived as positives.

I make that mistake myself all the time, What helps me avoid it is this: you want a number that is a property of the test method alone, independent of case distributions. A ratio between misidentified positives and identified positives (or true positives) would depend on sample distribution.

The metric you're talking about is called the "false discovery rate": https://en.wikipedia.org/wiki/False_discovery_rate

I also have a hard time keeping these all straight: https://en.wikipedia.org/wiki/Sensitivity_and_specificity#Ap...

50% accurate I'd agree is useless.

But the parent said 50% false positive, presumably with close to a 0% false negative would be VERY useful and save potentially millions of lives. We need enough tests yesterday or so to avoid a repeat of Italy and a 50% false positive rate (with a very low false negative rate) could help do that.

Even the properly-conducted version of this test has a fairly substantial false negative rate, somewhere around the 30% mark, and if it's done by students using samples that might not be taken correctly on a rigged-together testing setup that's going to get worse. Seriously, you might as well flip a coin.

A test with a 50% false positive rate, administered to 100 million people who aren't infected, would say 50 million people were infected.

False positive rate is a confusing term; it means the % of tests that should be negative that report a false positive.

I think the term for what you're probably thinking of (% of positive results that are false positives) is the false discovery rate.

That’s fair, if the assumption is that false negatives are very low or 0 (in which case it’s a super high accuracy test at low contamination rates and super useful).

And yeah obviously we desperately need a decent test like three months ago.

A test might have a high false positive rate, ie “low specificity”, but can still have “high sensitivity”.

That would mean that while you catch most cases of the virus, you’d also get a bunch of false positives. Flipping a coin as a hypothetical test would give false positives and negatives, or low specificity and low sensitivity.

Isn’t false negative specificity and false positive sensitivity?

https://en.wikipedia.org/wiki/Sensitivity_and_specificity

No, sensitivity is TPs / all positives (detected TP and FN). It’s the green half of the diagram in your link. The language is very easy to get tripped up by though. :)

I think I’m getting tripped by false positive, true positive.

A test with high sensitivity will have a low false positive rate.

A test with high specificity will have a low false negative.

So having a test with high sensitivity but low specificity will result in trust in the positives, but not trust in the negatives?

For me it’s easier to think of sensitivity as being “sensitive to the true positive” without saying much about false positive or negative.

A sensitive test will catch many positive cases. It may or may not have false positives though, eg a test that’s always gives the right answer vs a test that always returns positive no matter what.

A specific test will give you few positive results when the true answer is negative. You could use it to rule something out. One test might say “patient has A or B condition”, and a second test with high specificity may then rule out A or B, leaving B or A, respectively, as the probable condition.

I think you need to learn the difference between false positives and false negatives. A 50% false positive rate with no false negatives is absolutely better than 50% false negative and positive.

False negatives miss infected people and you fail to quarantine. False positives just mean you quarantine too many.

You’re not differentiating between sensitivity and specificity - in this case the test is highly sensitive but not specific. That means that it could be effectively hard to rule out cases while failing to provide strong evidence of actual infection in individual cases.

You're applying conventional knowledge to an unusual situation where it doesn't apply. The current test in my country is something like "was sitting within 4 rows of a known patient on a plane". That's obviously got a massive false positive rate but we do it anyway. The action you take when officially tested positive is the same as you take when learning you had contact with a patient - isolate yourself at home. There's no wasted drugs because there's no treatment for it! At worst, you might do an official test, but if those are in short supply, they're not going to be given to anyone who just walks in with a home-made result.

>A test needs to be materially more accurate than the odds of having the disease to be worth anything.

Shouldn't it just need to be better than 50/50?

>Classic thought experiment: if 0.0001% of the population has a disease and a test is 99% effective and you test positive, what are the odds you have the disease? Answer: 1%.

My odds still changed dramatically and the cost of a false positive is just me hanging out at home and not visiting my parents. I'm not getting a biopsy or taking expensive medicene.

Given one of the best pieces of advice about managing COVID-19 is to "Act Like You Already Have it" [1], a less accurate test with more false positives isn't necessarily a bad thing.

[1] https://www.huffingtonpost.com.au/entry/act-like-you-have-co...

Aren’t the primers supposed to be pretty selective for the RNA sequences being targeted? Otherwise any other random RNA in the sample (like from some other viral infection) would result in a false positive, right?

Once you have a successful PCR, you have billions of copies of your target sequence that are easy to aerosolize and/or transfer by contact onto your pipettes, gloves, and other lab surfaces. When a few of these molecules find their way into a new PCR you just set up, you get a false positive. There are protocols clinical labs employ to avoid this (e.g. never open the cycled PCRs in same room where you set up new PCRs, etc.) but they require strict adherence. (Or you can just give up all your sensitivity and only detect the cases with a huge viral load.)

Looking at the assay - it uses a dual labelled probe. That gives the specificity in addition to the PCR primers. Primers themselves presumably aren't selective enough.

> they're rife with false positives.

Is it possible that some of the positive cases of Corona are actually positive cases of Influenza?

No, it is not, see the numbers compared by the Italian National Biotechnology Association:

https://en.wikipedia.org/wiki/File:Is_COVID-19_like_a_flu%3F...

Each bar is one week.

Flu has totally different speed of increase and the amount of people needing treatment and dying.

We know what is the cause of the increase, the scenario repeats in many countries across the world, it's not one isolated setup.

I was curious about the possible mix up of flu and covid by the testing kit since they are similar, I am not denying that covid exists, just that if the tests are mixing the two up.

No, the covid testing kit was tested with more than a thousand probes and there wasn't a single false positive.

Source: Christian Drosten (he created the test) on the NDR Coronavirus Podcast

There is another source (in German) claiming the same, HN comment here: https://news.ycombinator.com/item?id=22627958 and source here: https://www.ndr.de/nachrichten/info/16-Coronavirus-Update-Wi...

> Regarding your question about if we might accidentally test for flu or other, already existing corona virii:

> This was a topic of discussion in yesterday's talk [1] with Dr. Drosten, a virologist who played an important part in the development of the currently used PCR test. He said that there were extensive studies done with hundreds of samples from both flu patients and patients infected with other corona virii and none returned a positive result. The only other positive results were from corona virii that are special to certain animals (bats, some cows IIRC), but none of those are present in humans. So the accuracy of our current PCR test for SARS-CoV-2 seems to be extremely high.

Edit: realize now that it's the same person (Dr. Drosten) so same source, but different format (podcast vs transcript)

> And theoretically, the test would give a positive result on some bat corona viruses, but they do not affect humans.

What if I have flu and that corona virus which do not affect humans. People will think I have COVID.

Edit:

The test developed was done against 75 people with various other diseases as listed below to find out the false positive rate. With the 75 samples (having listed diseases), the false positive rate was 0.

  Clinical samples with known viruses Number of

samples tested in all three assays HCoV-HKU1 2 HCoV-OC43 5 HCoV-NL63 5 HCoV-229E 5 MERS-CoV 5 Influenza A (H1N1/09) 6 Influenza A (H3N2) 5 Influenza A(H5N1) 1 Influenza B 3 Rhinovirus/Enterovirus 3 Respiratory syncytial virus (A/B) 6 Parainfluenza 1 virus 3 Parainfluenza 2 virus 3 Parainfluenza 3 virus 3 Parainfluenza A or -B virus 5 Human metapneumovirus 3 Adenovirus 3 Human Bocavirus 3 Legionella spp. 3 Mycoplasma spp. 3

   Total clinical samples 75

I hope we didn't any other major common virus, and also is 75 a big enough number to say that the false positive rate is definitely 0?

> What if I have flu and that corona virus which do not affect humans.

By definition, you can't have the virus which doesn't affect humans. The virus that causes Covid-19 is called SARS-CoV-2:

https://en.wikipedia.org/wiki/Severe_acute_respiratory_syndr...

2 means it's only a second ever recognized such. The first caused the original SARS in 2002-2003.

And that low chance of false positive can be easily derived (as in, just looking at the bars) from the Italian National Biotechnology Association graph, based on the known cases in Italy, which I've cited before. The numbers are orders of magnitude different, so the same can be concluded -- the can be no significant misrepresentation. There's the whole Italy and actual people dying or having to be connected to the machines to keep them alive -- a much bigger and more horrifying confirmation of low chance of false positives than just comparing thousand probes.

"mkagenius" sadly simply reiterates false claim dressed up as a restatement of the question after completely ignoring the answer to which he responds.

The viruses are not similar at all, the symptoms are to some degree.

SARS-CoV-2 is in the same family as SARSv1, MERS, a number of viruses that cause the common cold (229E, OC43, NL63), plus a number of bat viruses. But not influenza!

The test would cause a false positive for the most related ones, specifically SARSv1 and some bat viruses.

We really need a test for anti-bodies as well.

Antibody tests are also fairly unreliable in a number of circumstances as well.

The early CDC tests were not accurate, or the labs were not, correct? I am not qualified to speak to this, however, inaccurate results (positive and negative) could lead to overloading the healthcare system. https://www.technologyreview.com/s/615323/why-the-cdc-botche...

While this has been addressed by the CDC, how is bootleg version different to not cause the same issues?

It was only one of the tree parts of the CDC test that was giving false positives in most cases. Also, the experts on This Week In Virology thought that the CDC was being too clever in how it designed its test and that's why it got into trouble but I don't feel qualified to try to explain why exactly.

A false positive in the case of corona virus shouldn't be a problem since you shouldn't seek medical treatment unless the symptoms become severe.

As others have said, it's a huge problem if it leads people to think they're now immune to the coronavirus.

The steps are trivial to write out but executing this for someone inexperienced is not trivial. However, someone in the field, no doubt can do this, I'm sure.

My thoughts exactly. This is like saying it's trivial to do face recognition. All you have to do is load some libs, train the NN on them, and use the trained network on new data.

In practice everything from software versioning to knowing what might go wrong in NN training is gonna be a roadblock for someone who hasn't been shown.

As someone who had experience in both - PCR is similar, if not simpler to setting up NN. You just need equipment.

And setting up a NN is easy, you just need a laptop.

In general, if you find yourself using the phrase "you just need (to)..." Then you should probably take a step back and consider the actual work that goes into "just" doing the work.

Or, in the case you've outlined, not even doing the work, but just ordering the PCR equipment, since apparently the kit will just run itself? (I'm also assuming here that you realize that this isn't PCR + gel electrophoresis, but qPCR.)

The hard part is to do 10K tests per day like the South Koreans did to successfully contain the spread of covid-19 (or 60K tests per day since the USA have 6 times the Korean population).

This would include gathering saliva samples and bringing them to the PCR machine in a safe way.

New York State is at that level of testing per-capita, although a bit belatedly: it tested 7500 people between the Wed 3/18 and Thu 3/19 updates. If all states were testing at that per-capita rate, there would be about 125k tests per day nationwide.

I wonder if primers are in short supply? CDC authorized only two kits. Are the sequences known? Can you custom order/synthesize your own?

edit: sequences are apparently here https://www.cdc.gov/coronavirus/2019-ncov/downloads/rt-pcr-p...

If you put in an order for the primers by noon, you can have enough of them in your lab by the morning to run a thousand tests, maybe 10,000.

This comments reads like the classic "dropbox is trivial, it's just rsync" except it's more like "dropbox is trivial, it's just rsync on kubernetes with a reactJS frontend and a backend written in Haskell"

It's more like "We already have implemented dropbox for .jpg-files, also doing it for .png-files is trivial".

You won't be doing all this [PCR] from scratch at home McGyver-style (okay, maaaaaybe with a biochemical education) - but if you've got access to a biochemical lab that's doing RT-PCR anyway, has all the machines and all necessary chemicals (expect the COVID-specific markers) it's much more straightforward.

2d later, not that anyone still sees this. But: I know from a friend that their university lab is making plans to have their PCR-cabable equipment and staff on standby to run corona tests.

Maybe for someone with no bio background. But these are pretty standard techniques in molecular biology. The only tricky part is collecting the sample without exposing yourself to the virus.

Yeah, the Dropbox example doesn't make sense. Dropbox is hard to build because it needs to work at scale and for general enterprise purposes. Similarly, regional/state-sponsored testing is hard because it needs to work at scale and for a large population of people.

Running these kits out of your garage is like running a tweaked, existing file system for a home network. Not completely trivial, but it's not Dropbox, guys.

> Dropbox is hard to build because it needs to work at scale

And you don't think this test needs to scale, with the millions of people needing to be tested?

As far as I can read, this is literally the next sentence in my comment.

Whoops, so used to the "it's so easy" comments on here that I missed that part.

These are techniques high school students learn and practice in AP Bio... at least I did, 25 years ago.

Yes, and middle school students learn to write "Hello World!" HTML pages. So, by that logic, any web developer is interchangeable with a middle schooler.

Really? PCR was pretty new at that point.

Reading through some archived curricula, it was in a 1993 practice exam, at least. [0] I'm pretty surprised as well.

[0] http://www.csun.edu/science/biology/AP_biology/text%20files/...

no it isn't . there are lot of research labs that can do the test. the problem is safety, not the test itself.

...and to dig one layer deeper, the actual problem is diffusing blame in cases where the test is wrong or someone wants to blame the lab for the fact that they caught the bug, not safety. That's what certifications are for (in this case), and that's the real reason why running without one is risky.

risk tolerance changes in times of crisis though

Statutes of limitation run past the crisis when the lawsuits phrased, "you should have known" come out.

Yeah, imagine the janitor gets exposed to the virus, they then expose their entire family, those people then expose more, etc. You could easily kill someone.

> a band matching coronavirus positive control

Is the band matching the specific SARS-CoV-2 or it's just recognizing any coronavirus in circulation? AFAIK coronaviruses aren't that rare and are present in humans every year.

> Is the band matching the specific SARS-CoV-2 or it's just recognizing any coronavirus in circulation? AFAIK coronaviruses aren't that rare and are present in humans every year.

I've heard a podcast (in German) [0] with Christian Drosten who is one of the co-authors of the pt-pcr paper [1]. The relevant excerpt from the transcript (deepl translated):

> A very, very large validation study has been conducted. I would have to open it on my computer to go back to the numbers. But we have large numbers of real patient samples - with known positive evidence of other corona viruses and all the other cold viruses that we know of, and a whole number of these for each individual virus - and a whole number of patient samples, that is hundreds of samples with other corona viruses and other cold viruses, have been tested in this test. And not once has there been a false positive reaction. So this test does not react against any other human coronavirus or any other human cold virus.

> It is true, but this is of course completely misleading information, in theory this test would react against the old SARS corona virus. But that has not been present in humans for 16 years. And in theory, this test would also react against a whole range of bat coronaviruses, but they do not exist in humans either.

[0]: https://www.ndr.de/nachrichten/info/16-Coronavirus-Update-Wi...

[1]: https://www.eurosurveillance.org/content/10.2807/1560-7917.E...

the biggest fear is the handling of the samples and the possibility to get infected. Is there a way to make this completely automated? I know institutes in many countries have asked to perform PRC but the part of submitting samples en masse is the most critical, and labs need safety level certification for dangerous viruses. Drive-in submission is a solution, but sample depositories are even better

Would this be easy to do yourself if you were able to find (or make) a PCR machine?

FYI, the same core equipment and supplies will also allow you to do dna-based lab work, such as crispr/cas and fingerprinting for phylogeny, at home. If you're clever, you can replicate the same cutting-edge work step-by-step from published research papers and then use that knowledge to run your own experiments.

I'm currently working on adding the gene for bioilluminescence to p. cubensis fungi. If I can successfully breed offspring with the desire genotype, I hope to sell the novel spores and use those funds to invest in a variety of altruistic research like individualized DNA-based cancer treatment.

How hard is it to get started doing stuff like this? Do you need thousands of dollars of lab equipment or is the entry barrier relatively low? Making your own bioluminescent genetically modified organisms sounds really neat

You need a set of pipettes, a thermal cycler (i.e. https://www.ebay.com/b/PCR-Thermal-Cyclers/185231/bn_1153835... ), some consumables (i.e. pipette tips, vials, gels), reaction reagents (listed by the top level comment), a centrifuge, and probably some other things I'm forgetting. For some reactions you are much better off using a quantitative PCR machine but those are quite expensive.

If you just want to use molecular biology to make things glow see: https://www.minipcr.com/products/p51-molecular-glow-lab/

thanks so much, this will be a fun rabbit hole i think

Governments could open source all patents and design specifications for all relevant technology.

You can make your own pcr machine with thermometers, 3 pots of water on your stove, and a stop watch

You would also need a microcentrifuge and pipettes and stuff, but yes.

You need a quantitative real-time PCR machine, not just a PCR machine. Not cheap.

EDIT: You can't just put this on a gel. Something that comes up at 35 cycles (likely negative - needs re-running) looks exactly the same as something that comes up at 15 cycles (likely positive). The quantitative part is important.

2nd EDIT: it uses dual-labelled probe - end point PCR is absolutely not good enough.

Totally wrong. Take a sample out in 15 cycles. Take a sample in 30. Run a gel. Compare. Done. Most of the issues you think exist are because an RT pcr machine cannot distinguish between primer-dimer and pcr product. A gel can distinguish.

This argument is pointless as the assay relies on a dual-labelled probe. Look up how that works [1].

Anyway, an end-point PCR is not quantitative. You could do semi-quantitative on a gel, but it's a rubbish technique that was rightfully forgotten years ago, as it requires a lot of optimisation and has very little reproducibility.

[1] https://www.sigmaaldrich.com/technical-documents/articles/bi...

You seem to miss the point entirely. The point is that the assay doesn't NEED or REQUIRE dual labelled probes at all. The only need for quantification at all is to avoid mistaking signal for spurious noise (e.g., primer dimer). The gel provides both semi-quantification and provides amplicon size, which massively increases specificity of an end-point version of the assay. It is perfectly adequate.

The larger point is that there are many ways the test could have been designed around any possible combination of supply roadblocks (no DNA extraction reagent, choose phenol/chloroform; no cotton swabs, use paper; I could go on and on), so there are no excuses as to the massive and unwarranted and fatal delay in rolling out comprehensive testing in the U.S. Sure some work arounds are less desirable, but the alternative we have chosen is to fail a number (~>5,000) of people who didn't have to die by letting this thing spread without adequate testing.

you really think anyone cares how much of the corona you got in the particular blob of snot you gave for testing?

You deserve a 1000 upvotes for your succinct and accurate comment.

You are totally wrong. Your comment is dangerous misinformation. NO coronavirus assay requires quantitation.

Or you could just run the PCR product on a gel for much cheaper...

Edit: A gel gives you the ability to select for size as well, which a qpcr machine does not. Not sure why I have to reply in the form of edits.

on a gel you see the size, something coming up with exactly the right size even at 35 cycles indicates that there was at least some of what you designed the primers on in the sample. if you want higher certainty, with three primers you can get two different sizes ruling out any doubt.

with the rtpcr machine readout which only gives you the signal corresponding to how much was amplified over time you need tricks like the dual labelling.

Why is that? Is it really impossible to do the test on a simple PCR machine, or is it just too slow?

pPCR is quantitative - measures fluorescence. End point PCR just measures whether you have the sequence there or not. It's slower, more work, and prone to false positives (see edit above).

I saw a whole kit including laptop, software, and license for $3,500 buy-it-now on eBay last week.

Yes. You can buy a PCR machine for about $1,000 US. You can build your own agarose gel box for about $100. THe power supply might be tricky, but probably not. The CDC has published all of the primer and probe sequences. It is trivial to run the assay yourself if you buy the PCR machine and a few water baths for the RT and DNAse reactions.

If you aren't worried about safety then you can use e.g. a full bridge rectifier hooked up to 110 V AC to get a usable (but shitty) agarose gel power supply. I really don't recommend this but it does show how easy it can be.

For someone who has zero previous knowledge, what are some good online resource to learn this stuff? Is it realistic to do in, say, a couple of weeks?

it's a specialized swab shortage

You ruined the Clickbait \s

100+ disappointments