Dear Neda,

I don't know about wavelenght diapason of your laser but many years ago I studied the effect of the short exposition to visible light of the different wavelength on cultured cells (fibroblasts). In brief, the green light stimulated cellular metabolism and proliferation, the blue light inhibited proliferation without the ihhibition of mitochondrial respiratory enzymes (MTT) and the red light stimulated proliferation but destabilized cellular membrane (LDH leakage) so the green light could be classified as stimulating, blue light - as stabilizing and red light - as irritating. Unfortunately, these results were not published separately but only included in my doctoral thesis (in Russian).

However I can cite some article(s) where the low-intensive helium-neon laser (630 nm or 660 nm; 2,4-4 J/cm2) stimulated cellular proliferation: Webb C., Dyson M., Lewis W.H.P. Laser Surg. and Med., 1998, 22(5), 294-301; Ben-Dov N., Shefer G., Irinichev A. et al. Biochim. Biophys. Acta Molec. Cell. Res., 1999, 1448(3), 372-380.

Hi Neda,

I am not aware of the confluency for your cell types. Is it possible that your cells were confluent (or any other reason for growth inhibition) before irradiation and hence had attained stationary phase, but the irradiation killed some cells and paved way for the other cells to restart proliferation?

If you are using the MTT-assay imediately after treatment, you are most likely not assaying proliferation. I suggest you look in to the biochemistry of the MTT assay, which depends on (mitochondrial) dehydrogenase activitiy (see e.g. Berridge, M.V., 1993). The laser treatment may be producing rapid effects at the membrane level.

It is not clear wavelength of laser you are using for your studies.If you are investigating the effect of lasers on proliferation/growth of cells, I suggest you to do MTT assay after 24h of exposure to laser as manifestation of effect on growth require time. Also, since photobiostimulatory effect of laser depend on optical parameters, effect of irradiance (mW/cm2) and light dose (J/cm2) need to be studied.

Lasers of wavelength ranging 630-680 nm(red) and 800-1100 nm (NIR) have been shown to induce biostimulatory effects i.e. proliferation and differentiation of different types of cells including keratinocytes, fibroblasts, endothelial cells, myoblasts, and osteoblasts. Some of the references are 1. IEEE J Sel Top Quantum Electron. 2016; 22(3),

2. Photomed Laser Surg. 2010;28 Suppl 1:S151-6.,

3. J Physiol Pharmacol. 2013;64(3):387-91. 

Check step by step,

Next time do uniform seeding of cells (5000 cells/ well in 100uL media) by counting cells, use trypan blue. Also MTT is done in 96 well plate, I am not sure whether you seeded cells prior or performed your MTT in 6 well plate.

Read plate at 595nm.

Most importnat do not incubate cells for more 24 hours prior any treatment or even laser.

Might be in 2 days (48 hours), cells confluency is high so there is not much difference in result.

Dear Dr.Mikhail Yurievich Eropkin

I am using the 1064 nm laser operating for both CW and short pulse laser. (green light).

I apply the laser on cells (96-well-plate)for 50 seconds. Then I did MTT Assay 0,6, and 24 hours after treatment by laser. The color didnt change. And my control is lower than my result after laser. I seeded 8000 fibroblast cells/well.