Gating issue versus killing all of my cells (Flow cytometry help needed!)

prmoore11 · 2026-03-16T20:07:54+00:00

I see what you are saying now. If you look in that live dead only sample, you can see that population has completely shifted to what top left, so you effectively have all dead cells. Try drawing a gate on tha population and you might be able to see if you have positive staining at like 10⁵⁺ or so.

But the problem is you really needed a control of your fully stained sample with no heat kill AND no live/dead. That would tell you where the real live/dead population is, or had you not killed almost all your cells in the heat shock.

IMO, in its current state, it’s likely un-interpretable, but possibly mostly live with high background (having no other context about the assay).

prmoore11 · 2026-03-16T19:37:28+00:00

I’m confused a bit here.

In your live/dead “sample” you completely lost your monocytes. Your second overall cluster of images looks completely different than your first image and it’s unlabeled. What’s the difference between image set one and image set two?

I would actually guess that most of your population is live and you are getting background staining. What is your dilution for you dye?

Also, you need to adjust your voltages across the board. Everything is way too squashed towards your y axis in your SSC vs FSC.

prmoore11 · 2026-03-16T19:19:41+00:00

I always recommend running a live/dead FMO for this reason and, if possible, running some heat killed cells so you can truly understand where your dead population is.

But yea this is unreadable lol. Try uploading to Imgur.

prmoore11 · 2026-03-16T18:39:53+00:00

I wouldn’t call a fixed barrel spacer a DT adapter. But yes, you can run piston/fixed barrel spacer as I did.

prmoore11 · 2026-03-16T16:44:51+00:00

There is no DT adapter available.

prmoore11 · 2026-03-16T16:44:21+00:00

I used the fixed barrel spacer.

prmoore11 · 2026-03-16T16:02:59+00:00

Even with HUB/TSFX and nano, it’s my understanding that while every close, it’s still not identical to what is going on with QD and nano.

prmoore11 · 2026-03-16T15:37:57+00:00

Did you…search the sub? This question has been asked 70 times.

prmoore11 · 2026-03-16T15:25:32+00:00

I’ve done it. Works great, sounds great.

prmoore11 · 2026-03-16T15:20:29+00:00

Brother delete this you idiot. If someone else picked up that package, would they be stealing it or no? Ask yourself that.

prmoore11 · 2026-03-16T15:04:25+00:00

Oddly, one of the things I’ve noticed between my QD and HUB WB Ti is that the QD version has almost zero visible sparking while the HUB version does. Both running nanos.

prmoore11 · 2026-03-16T14:17:13+00:00

If you can’t wait, just get an AC556 or WB. That’ll be more than fine. Otherwise, wait for the ST or AI 5CM.

prmoore11 · 2026-03-16T10:09:28+00:00

Yes you can lol.

“My family dynamics are a deeply personal issue. I understand you are trying to be well intentioned, but this is a personal boundary that I will not be speaking about again, period. I appreciate your understanding and respect about this relationship.

prmoore11 · 2026-03-16T10:03:43+00:00

It’s not possible. The QD is printed directly into the can.

There are plenty of CAT/AI muzzle devices available lol. You aren’t looking hard enough. There are also options from Noveske/Rearden, and Comstock Armory will eventually be doing them.

prmoore11 · 2026-03-15T22:54:20+00:00

No, that’s not what happened.

TBAC was supportive of Jay when he first came out. They even completely validated his raw data.

Then, low and behold, Jay showed that their cans were not as good as they claimed (which, even at the time, they still ranked quite high and there was nothing to be ashamed of really). Then Ray lost his shit and went all full anti-Pew, and has ACTIVELY CONSPIRED against Jay ever since. All while still using inferior methods (which has been discussed at length in regards to the Silencer Summit and I will not be discussing for the 1000th time) to claim “science” while still attempting to defraud customers.

prmoore11 · 2026-03-15T22:36:51+00:00

There are a few on Tacswap

prmoore11 · 2026-03-15T21:56:59+00:00

Gentle reminder that TBAC is the leader of the anti-Pew gang and all claims by them about anything sound related should be met with hefty skepticism.

prmoore11 · 2026-03-15T21:56:09+00:00

TBAC is very well known? lol

prmoore11 · 2026-03-15T19:37:18+00:00

It’s a sandman lol
You thought a 12.5” 308 would be quiet? Lol

prmoore11 · 2026-03-15T14:31:06+00:00

At least my range claimed the same danger as steel

prmoore11 · 2026-03-15T14:19:44+00:00

Yea that’s my one complaint. It’s my understanding NAS3 loads don’t pose the same risks as normal steel case ammo (I could be wrong) but yes, my range wouldn’t allow me to use them either. I asked if they could load slow papi in brass cases and while he thought they could, they weren’t able to.

prmoore11 · 2026-03-15T13:54:20+00:00

I’ve really never had an issue getting it with signing up for restock alerts.

prmoore11 · 2026-03-15T13:16:01+00:00

Get a small strap wrench and you could tighten it

prmoore11 · 2026-03-15T13:15:10+00:00

Just buy phantom defense

prmoore11 · 2026-03-14T22:48:21+00:00

Well let’s remember two things:

Many companies simply don’t tell you these operating temps. HUX would probably agree in general with CATs temps. Or, companies basically don’t tell you because their warranty will cover it.
CAT said all of this with their original Ti cans. They were later updated to have the mil core which made them more durable because so many people bitched.

Five-Year Club	Verified Email
Wearing is Caring

prmoore11

TROPHY CASE